The implications of shedder status and background DNA on direct and secondary transfer in an attack scenario

In court questions are often raised related to how trace DNA was deposited, directly during the crime or innocently for instance by secondary transfer. It is therefore of interest to have knowledge of the probability of transfer or secondary transfer in different situations. Factors that could influence transfer probabilities are background DNA and the shedder status of the involved persons. In this study, we have classified participants as high or low DNA shedders. We observed DNA transfer in a simulated attack scenario, and demonstrated that shedder status has a significant influence of transfer rates. We have examined the background DNA in samples from T-shirts worn in an area with frequent human traffic and detected multiple contributors. We further demonstrated that DNA from co-workers of a T-shirt wearer can be secondarily transferred from the environment and detected in samples, and that the composition of background DNA is correlated with the shedder status of the wearer. Finally, we have illustrated the inference with the results of transfer probabilities and a fictive case with the use of a Bayesian network.     

Collaborative swab performance comparison and the impact of sampling solution volumes on DNA recovery

The collection of DNA traces marks the first step determining the success of genetic analysis. This study aimed to identify and validate a suitable alternative to the currently used ForensiX Evidence Collection Kit containing a cardboard box for swab storage. This box has to be folded at the crime scene, which is time-consuming and carries the risk of potential contamination and handling difficulties.A collaboration study involving three police departments and one laboratory for forensic genetics was performed to compare the currently used swab against three challenger swabs: ForensiX SafeDry, Copan 4N6FLOQSwab™ Genetics and Copan 4N6FLOQSwab™ Crime Scene. Mock samples consisted mainly of touch DNA, but also blood, saliva and semen were applied to twelve items with different surfaces. Every organisation contributed with three DNA collectors, whose individual collection efficiencies were investigated. The challenge of preparing homogenous traces, especially touch DNA, was addressed by enhancing hand contact frequency and sampling area. As a further part of the swab comparison study, we describe for the first time the influence of different swabbing solution volumes on the sampling efficiency of the different swabs.The application of touch DNA was also tested for a further swab type, the Sarstedt Forensic Swab, which yielded such low DNA concentrations that it was excluded from the collaboration study. The Copan Genetics and Copan Crime Scene swabs yielded significantly lower DNA concentrations than the currently used ForensiX Evidence Collection Kit and ForensiX SafeDry swab. The inter-individual performance results of the operators revealed significant differences in sampling skills. Comparing different swabbing solution volumes showed higher DNA yields or no significant difference for the ForensiX Evidence Collection Kit and ForensiX SafeDry than the Copan Genetics, depending on the item or trace type swabbed.Our results highlight the importance of validating first-step components that are decisive to the success of DNA typing in the context of specific sampling procedures and laboratory methods. Also, the significance of individuals' securing variations, principally unknown for crime scene investigation and laboratory teams, is emphasised for the first time, offering a practical approach for improving and training DNA collecting activities and ensuring the optimal securing evidence process. These findings increase the knowledge of impacts on DNA collection and, thus, benefit other laboratories and forensic services, particularly when using the same extraction methods.     

Trace DNA evidence dynamics: An investigation into the deposition and persistence of directly- and indirectly-transferred DNA on regularly-used knives

Empirical data on the transfer and persistence of trace DNA are crucial to the evaluation of forensic DNA evidence. This evaluation can be complicated by the occurrence of indirect DNA transfer; the possibility of which is well established, but research into such transfer is often focussed on unrealistic situations, e.g. handling of DNA-free items after participants have shaken hands for 1–2 min. To simulate more realistic scenarios, this study investigated the deposition and persistence of both directly- and indirectly-transferred DNA on knives that had been artificially set up as ‘regularly-used’. Each knife was handled in a prescribed manner by a specific participant over two consecutive days to simulate regular use. Each participant then shook hands for 10 s with a fellow volunteer and immediately stabbed one of their knives into a foam block repeatedly for 60 s. DNA was recovered by mini-taping from triplicate sets of knife handles from four pairings of volunteers after regular use, and at one hour, one day and one week after the handshaking and stabbing events.Total amounts of DNA recovered from the knives, regularly used by a single person, varied among individuals; one volunteer consistently deposited significantly greater amounts than the others, whilst another volunteer did not always leave complete profiles. DNA attributed to the regular user persisted for at least a week, declining with increasing time between DNA deposition and recovery. Non-donor DNA was co-deposited at <5% of the profiles recovered, except for one volunteer, who consistently left DNA from their romantic partner on their knives at ∼25% and ∼11% of the profiles before and after the handshaking and stabbing events, respectively. In three pairings of volunteers, after the handshaking and stabbing events, alleles that could be attributed to the respective handshakers’ profiles were detected as partial minor profiles, equating to ∼10% of the profiles recovered. For the fourth pairing of volunteers, only complete single-source DNA profiles matching the regular user’s profile were recovered. However, it is important to note that, when indirectly-transferred handshaker DNA was detected, it declined with increasing time between DNA deposition and recovery.These data provide an initial insight into the detection and persistence of directly- and indirectly-transferred DNA that extend the data already available on forensic DNA transfer. The results herein suggest that the sooner an item is sampled after an offence has occurred, the greater the chance of recovering indirectly-transferred DNA, which has implications for forensic reconstructions.     

Statistical and population genetics issues of two Hungarian datasets from the aspect of DNA evidence interpretation

When the DNA profile from a crime-scene matches that of a suspect, the weight of DNA evidence depends on the unbiased estimation of the match probability of the profiles. For this reason, it is required to establish and expand the databases that reflect the actual allele frequencies in the population applied. 21,473 complete DNA profiles from Databank samples were used to establish the allele frequency database to represent the population of Hungarian suspects. We used fifteen STR loci (PowerPlex ESI16) including five, new ESS loci. The aim was to calculate the statistical, forensic efficiency parameters for the Databank samples and compare the newly detected data to the earlier report. The population substructure caused by relatedness may influence the frequency of profiles estimated. As our Databank profiles were considered non-random samples, possible relationships between the suspects can be assumed. Therefore, population inbreeding effect was estimated using the FIS calculation. The overall inbreeding parameter was found to be 0.0106. Furthermore, we tested the impact of the two allele frequency datasets on 101 randomly chosen STR profiles, including full and partial profiles. The 95% confidence interval estimates for the profile frequencies (pM) resulted in a tighter range when we used the new dataset compared to the previously published ones. We found that the FIS had less effect on frequency values in the 21,473 samples than the application of minimum allele frequency. No genetic substructure was detected by STRUCTURE analysis. Due to the low level of inbreeding effect and the high number of samples, the new dataset provides unbiased and precise estimates of LR for statistical interpretation of forensic casework and allows us to use lower allele frequencies.     

Touch DNA: The effect of the deposition pressure on the quality of latent fingermarks and STR profiles

Latent fingermarks (FMs) present unique, and sometimes the only, evidence found at a crime scene. Several factors affect their quality, including deposition pressure (DP). Its effect on FM size and quality, and on STR amplification success rate, is an emerging area of interest in forensic science. This study examined 540 FM samples, each consisting of index, middle and ring fingers, deposited by 30 donors on glass, polythene (PE) and paper under a range of weights from 0.1 to 10 kg. Both length and width of FMs increased with the increasing DP. FMs deposited under lower (≤0.5 kg) DPs varied in size (p < 0.01), while those deposited at higher (≥3 kg) DPs were more consistent. FM quality on glass and PE, as determined by the AFIS minutiae count and by a fingerprint examiner on a scale from 0 to 4, improved with the increasing DP, but it deteriorated on PE at DP of 10 kg. FM quality on paper continued to improve from DP of 1 kg up to the maximum DP of 10 kg. The effect DP has on the efficacy of DNA profiling from latent FMs was significant as shown by an increase in the DNA amount recovered, the number of amplified loci per FM sample, and the number of forensically useful DNA profiles (defined here as those with ≥8 full STR loci detected) as DP increased. This effect was most pronounced with PE (R = 0.98) and paper (R = 0.96). Altogether, the success rate of DNA profiling varied from 16.3% in FMs deposited on paper to 21.2% and 22.5% of those on PE and glass. The highest number of useful DNA profiles was obtained from glass under DP of 10 kg. Forensically useful FMs obtained at low (≤1 kg) DP from all three substrates significantly outnumbered that of STR profiles, while an opposite, though less pronounced trend, was observed at high (≥3 kg) DP on PE and paper. Application of the simple device for collecting of FMs under controlled pressure designed for this study, and the palm-up mode of FM deposition as described, allowed us to eliminate the undesirable effect of the hand self-weight and to objectively assess the actual effect of increasing DP on FM size and quality, as well as on the efficacy of DNA profiling.     

DNA on drugs (part 2): An extended study into the transfer and persistence of DNA onto illicit drug capsules using realistic scenarios

Capsules are now the main form of ecstasy rather than tablets in Australia and therefore their examination is of interest to forensic drug chemists in Australia and possibly elsewhere. Recently, we used controlled experimental conditions to show that capsules may be a source of DNA that can be used to identify those involved in production and distribution of illicit drugs. The question remains: in realistic scenarios where there are more unknowns, can we still detect DNA, and determine whose it is, on the exterior of capsules? The concept of comprehensive forensic intelligence and investigations – utilizing both biological and chemical signatures – relating to illicit drug preparations (i.e., the capsules and their contents) may be of great use to law enforcement. Experiments were conducted with both semi-realistic and realistic scenarios where two volunteers were asked to firstly use an encapsulator and mimic the loading of capsules, then Volunteer 1 would count out the capsules that Volunteer 2 prepared, and vice versa. This was to simulate the scenario where one person was involved in the assembly of the capsules which were then separated into smaller bags of 10 capsules by a second person for distribution. Gelatine and vegetable capsules were tested, with 10 replicates used per capsule type, scenario, and volunteer (total n = 80 capsules). Volunteer 2 was included as a contributor to the DNA profiles generated from 100% of samples handled by them within the semi-realistic scenario, whereas the other volunteer could be included as a contributor in 65% of samples. For the realistic scenario, profiles could be generated with the inclusion of both volunteers as profile contributors in 15% of samples and from just one of the volunteers in a further 50% of samples (therefore in total, either both or one of the volunteers were detected in 65% of realistic samples). Surprisingly, it was not necessarily the case that the last person to handle the capsule was the major or only contributor. The potential variability in the DNA quantities that could be deposited onto the capsules of genuine illicit drugs is high and would vary on a case-by-case basis. Nevertheless, this study has indicated that in realistic scenarios where two people are involved in the later stages of illicit drug capsule preparation, that either one or both individuals may be identified, potentially opening new investigative leads for law enforcement agencies as well as offering new information for intelligence-led policing.     

Impact of SNP microarray analysis of compromised DNA on kinship classification success in the context of investigative genetic genealogy

Single nucleotide polymorphism (SNP) data generated with microarray technologies have been used to solve murder cases via investigative leads obtained from identifying relatives of the unknown perpetrator included in accessible genomic databases, an approach referred to as investigative genetic genealogy (IGG). However, SNP microarrays were developed for relatively high input DNA quantity and quality, while DNA typically obtainable from crime scene stains is of low DNA quantity and quality, and SNP microarray data obtained from compromised DNA are largely missing. By applying the Illumina Global Screening Array (GSA) to 264 DNA samples with systematically altered quantity and quality, we empirically tested the impact of SNP microarray analysis of compromised DNA on kinship classification success, as relevant in IGG. Reference data from manufacturer-recommended input DNA quality and quantity were used to estimate genotype accuracy in the compromised DNA samples and for simulating data of different degree relatives. Although stepwise decrease of input DNA amount from 200 ng to 6.25 pg led to decreased SNP call rates and increased genotyping errors, kinship classification success did not decrease down to 250 pg for siblings and 1st cousins, 1 ng for 2nd cousins, while at 25 pg and below kinship classification success was zero. Stepwise decrease of input DNA quality via increased DNA fragmentation resulted in the decrease of genotyping accuracy as well as kinship classification success, which went down to zero at the average DNA fragment size of 150 base pairs. Combining decreased DNA quantity and quality in mock casework and skeletal samples further highlighted possibilities and limitations. Overall, GSA analysis achieved maximal kinship classification success from 800 to 200 times lower input DNA quantities than manufacturer-recommended, although DNA quality plays a key role too, while compromised DNA produced false negative kinship classifications rather than false positive ones.     

Assessment of the transfer,persistence, prevalence and recovery of DNA traces from clothing: An inter-laboratory study on worn upper garments

Among the various items recovered from crime scenes or persons involved in a crime event, clothing items are commonly encountered and submitted for forensic DNA sampling. Depending on the case circumstances and the activity-of-interest, sampling of the garment may concentrate on collecting DNA from the wearer, or from one or more offenders who have allegedly contacted the item and/or wearer. Relative to the targeted DNA, background DNA already residing on the item from previous contacts, or transferred during or after the crime event, may also be collected during sampling and observed in the resultant DNA profile. Given our limited understanding of how, and from where, background DNA is derived on clothing, research on the transfer, persistence, prevalence, and recovery (TPPR) of DNA traces from upper garments was conducted by four laboratories. Samples were collected from several areas of two garments, each worn on separate working or non-working days and individually owned by four individuals from each of the four laboratories, and processed from DNA extraction through to profiling. Questionnaires documented activities relating to the garment prior to and during wearing, and reference profiles were obtained from the wearer and their close associates identified in the questionnaire. Among the 448 profiles generated, variation in the DNA quantity, composition of the profiles, and inclusion/exclusion of the wearer and their close associates was observed among the collaborating laboratories, participants, garments worn on different occasions, and garment areas sampled.