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DNA transfer: DNA acquired by gloves during casework examinations
Accepted Manuscript Title: DNA transfer: DNA acquired by gloves during casework examinations Authors: Mariya Goray, Erin Pirie, Roland A.H. van Oorschot PII: DOI: Reference:
S1872-4973(18)30428-9 https://doi.org/10.1016/j.fsigen.2018.10.018 FSIGEN 1990
To appear in:
Forensic Science International: Genetics
Received date: Revised date: Accepted date:
31 July 2018 26 October 2018 31 October 2018
Please cite this article as: Goray M, Pirie E, van Oorschot RAH, DNA transfer: DNA acquired by gloves during casework examinations, Forensic Science International: Genetics (2018), https://doi.org/10.1016/j.fsigen.2018.10.018 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
DNA transfer: DNA acquired by gloves during casework examinations Mariya Goraya,*, Erin Piriea and Roland A.H. van Oorschotb,c
Biometric Division, Victoria Police Forensic Services Department, Macleod, Australia
b
Office of the Chief Forensic Scientist, Victoria Police Forensic Services Department,
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a
c
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Macleod, Australia
School of Molecular Sciences, College of Science, Health and Engineering, La Trobe
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University, Bundoora, Australia
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*Corresponding author: Telephone: +61394509715
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Email: [email protected]
Highlights
Examiner variation in exhibit areas touched and timing of glove changes
Exhibit and staff derived DNA acquired by gloves during examinations
Risk of removal, addition and redistribution of DNA by gloves during examination
Examiners need to be cognisant of areas touched and timing of glove changes
Stages of removing exhibit from packaging and repackaging require special attention
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1. Introduction
DNA transfer prior, during and post criminal activity has been a topic of conversation among
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forensic scientists for some time now [1-38]. Questions relating to when and how the DNA material was deposited onto surfaces of interest are becoming paramount [39-41]. These
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questions are pertinent to the events relating to criminal activity as well as the activities of
investigators post criminal event. Contamination post criminal event by investigators have
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been observed [42-44] and can occur through various means, including: non- or inappropriate
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use of personal protective equipment when near and/or handling exhibits to be examined;
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contact of such exhibits with dirty tools and/or equipment [2, 45-52]. Here we focus on
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elements relating to the potential of DNA transfer via gloves during examination of exhibits
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by examiners within a forensic biology laboratory.
Most surfaces and objects that are regularly contacted will have some level of background
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DNA [22, 53-54]. The quality and quantity of this DNA will depend on the frequency with
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which these surfaces are touched/contacted, by whom/what and the type and regularity of cleaning involved [48, 52, 54-56]. Environmental DNA studies of forensic biology laboratories have demonstrated that surface, equipment and tools within the laboratory can
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also contain detectable quantities of DNA, that may have been derived from examiners, others that have entered the area or handled the packaging, equipment, tools, consumables, or from exhibits previously examined [48,52,52,54]. Unclean surfaces, equipment and tools within a forensic laboratory pose a potential risk in respect to contamination of exhibits to be examined.
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Contamination of exhibits can occur at any stage of an investigation. Gloves worn during examination are a key risk factor for such contamination. This risk will be dependent on how gloves are put on, what they touch and when they are replaced. Addition, loss and/or redistribution of DNA to/from/within an exhibit through handling during examination can
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have significant implications for the investigation, placing doubt on the meaning of any
results obtained [15, 37-48, 50-52]. Several studies have investigated potential DNA transfer
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during mock “casework” examination (for example with tools), allowing for some inferences of what can be expected in real casework. However, to our knowledge, no studies have
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investigated DNA transfer via gloves during actual casework examination.
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The aim of this study is to investigate the risk of DNA loss, addition and redistribution via
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gloves during real casework examination, as well as the appropriateness and frequency of glove changes by the examiners. Several examiners were video recorded during examination
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of a range of casework exhibits. The timing of all glove changes and all contacts made by each glove were documented. Each glove used during the examination was sampled at the
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moment of the intended glove removal. The profiles generated from these gloves were examined in combination with the profiles generated from the exhibits, persons relevant to
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the case, the examiner, the researchers and other staff.
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2. Materials and Methods
2.1
Experimental set up
New gloves (“InControl” long cuff nitrile (power free) gloves) were sampled (as negative controls) and placed in dedicated glove boxes with each six gloves separated by blotting
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paper, ready for use. The glove box openings were widened in order to minimize examiner contacts with the box and the gloves. The boxes were placed into new clip seal bags to minimize exposure until required. The unused gloves remaining within a blotting paper section were discarded after each examination. Four examiners (A-D) were each videorecorded during exhibit examination and each pair of gloves used by the examiners during
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examination starting with the pair used to open the exhibit packaging and finishing with the pair used to return the exhibit back to its packaging were separately sampled (left and right).
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The palm and fingers (palm side only and avoiding the cuff area) of each glove were sampled
by a researcher whilst still being worn by the examiner and the samples then processed as per
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case work protocols.
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Eleven separate examinations were conducted (5 by examiner A, 2 by examiner B, 2 by
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examiner C and 2 by examiner D). Each examination was of a single exhibit. Of these
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exhibits, 5 were examined for trace (metal pole, section of mattress, jeans, jacket, and bottle opener) and 6 for blood (jacket, hockey stick, two singlets, knife and t-shirt). Overall, 96
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pairs of gloves from the examination of 11 exhibits were analysed.
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The number, duration, sequence and types of contracts made by each glove during
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examination were documented from the video recordings and assessed against the DNA profiles obtained from the gloves as well as those obtained from samples collected from the
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exhibits by the examiners.
2.2
Sample Processing
Each glove was swabbed separately (wet and dry swabs) (150C swabs, Copan, USA) and the two swabs from each glove were placed directly into a 2ml tube for further processing. DNA was extracted, quantified and amplified as per manufacturer recommendations, using DNA
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IQTM (Promega), Quantifiler®Trio (Applied Biosystems) and PowerPlex® 21 (Promega) kits, respectively. Amplified products were run on a 3500xL Genetic Analyser (Applied Biosystems) and typed using GeneMapperTM IDx Software (Applied Biosystems) with a baseline threshold of 175 RFU.
Data Interpretation and Statistical Analysis
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2.3
The total DNA retrieved from each glove was calculated by multiplying the DNA
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concentration, as determined during quantification, by the volume of the extract (60 µl). The total number of alleles for each sample was determined by counting the total number of
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alleles detected at each locus (threshold: 175 RFU), excluding peaks that were determined to
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be potential artefacts (e.g. stutter) as per the laboratory interpretation method and alleles at
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Amelogenin locus.
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The minimum number of contributors was determined for each of the profiles obtained, as per our laboratory’s casework DNA interpretation method, using a modified allele count
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method which takes into consideration stochastic effects. Profiles were then interpreted and
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statistically evaluated using continuous probabilistic software STRmixTM (version 2.06) [5758]. This software weights genotype combinations and allows comparison to persons of
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interest (POI) and the staff elimination database, expressed as likelihood ratios. Furthermore, STRmixTM interpretation output provides additional information such as; the mixture proportions expressed as percentages. The comparison to the staff elimination database via
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STRmix involves calculating a Likelihood Ratio to each person on the database; without the use of theta.
The Kruskal-Wallis one-way analysis of variance was used to test if the k independent samples were from the same or different populations (at p<0.05 significance level). Given
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the continuous nature of the variables examined, statistically significant groups were analysed with the Mann-Whitney U-test and the relationship between the continuous variables using Spearman’s correlation (at p<0.05 significance level). Pearson’s correlation was used to measure the strength and direction of association that exists between two variables. Statistical
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analyses were performed in SPSS Statistics (IBM).
Quality Controls and Cleaning Protocols
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The laboratory space and tools were cleaned as per laboratory protocols with 1%
Hypochlorite followed by distilled water. Of all the control swabs taken from gloves used by
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the examiners prior to them being worn, forty-nine (chosen based on the DNA result from the
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gloves) were processed to check for possible contamination. Twenty-four control swabs from
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the gloves where staff were not excluded as contributors to the DNA obtained were chosen
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for analysis. The remaining twenty-five controls processed were chosen randomly. Both positive and negative controls were implemented during DNA processing steps as per
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3. Results
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casework methods and produced expected results.
3.1
General
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Figure 1 is an illustrative snapshot of some of the data collected during one of the observed examinations.
Ninety-five gloves were used to examine six blood exhibits and ninety-eight gloves to examine five trace exhibits. The number of glove changes per exhibit ranged from 4 to 15
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pairs (average 9). On average 10 pairs of gloves were used during blood exhibits examination and 8 during trace exhibit examinations. Total DNA detected on the gloves ranged from 0 to 2.26ng (average 0.21ng) with an average of 0.321ng detected on the blood exhibit gloves and 0.106ng detected on the trace exhibit gloves. Profiles were produced from 98% of blood exhibit gloves and 87% of trace exhibit gloves. The remaining 2% and 23% respectively
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(total 25 gloves) did not contain sufficient DNA to produce a DNA profile. The total number of alleles ranged from 0 to 121 (average 26). There was a statistically significant positive
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correlation between the number of alleles and total DNA detected. The number of contributors on the gloves ranged from 0 to 6 people (average 1.8). The average number of
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contributors on blood and trace exhibit gloves was 2.3 and 1.3, respectively. In general, the
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gloves used at the beginning and those used at the end of an examination had approximately
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twice as much DNA and number of contributors than other gloves.
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The total quantity of DNA, total number of alleles, and the number of contributors detected on the gloves used to examine blood exhibits were significantly greater than on the gloves
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used to examine trace exhibits. There were no significant differences in the total quantity of
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DNA, the total number of alleles, and the number of contributors detected between the left
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and right hand gloves.
When comparing the glove changes between examiners, three examiners had an average of 11 glove changes for blood and trace exhibits combined, and one examiner had an average of
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6 changes per examination. There were significant differences in total DNA quantities, total number of alleles and number of contributors detected on the gloves among the four examiners. Two examiners consistently had significantly more DNA, more alleles and higher number of contributors detected on the gloves when compared to the other two examiners.
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On average, there were six glove changes for soft/flexible exhibits including clothing and piece of mattress and ten glove changes for hard/solid exhibits including weapons, metal pole etc. There were no significant differences in total DNA quantities, total number of alleles and number of contributors detected on the gloves between the soft/flexible and hard/solid
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exhibits.
Of the gloves with alleles above the detection threshold 78% (131 of 170 gloves) had at least
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one unknown allele detected that could not be attributed to DNA reference samples relevant
to the case, the evidentiary profiles obtained from the exhibit or to the examiner. Sixty five percent of the gloves with unknown DNA alleles were from gloves used to examine blood
Comparisons with evidentiary, reference, staff and examiner profiles
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3.2
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exhibits.
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The profiles generated from the gloves were compared to the profiles obtained from the evidentiary exhibits, the DNA reference samples relevant to each exhibit and case as well as
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profiles of the examiners and those on the staff elimination database.
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In 54% and 27% of blood and trace exhibit gloves respectively the POI relevant to the case
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(either suspects and/or complainants) was not excluded as contributor(s) to the DNA results obtained. Complainant was POI on weapons and suspect’s clothing; while suspect was POI on the complainant’s clothing and weapons. No POI DNA was detected on two (metal pole,
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section of mattress) of the five trace exhibits; for the remaining three exhibits (jeans, jacket, bottle opener), the POI was not excluded in 1/28, 18/30 and 3/16 gloves respectively. In most instances where the POI was not excluded from the gloves, they were also not excluded from the evidentiary profiles generated during exhibit sampling. Interestingly, for one of the
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exhibits, a bottle opener, the suspect was not excluded, albeit with low support, (LR=22, with theta) from the gloves, yet was excluded from the evidentiary profile.
Comparison of the results generated from the gloves and exhibits showed that in most instances higher total amounts of DNA were detected in the evidentiary samples compared to
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the examination gloves (for both blood and trace gloves). Further, while 33% of the gloves had higher number of contributors compared to the corresponding evidentiary profiles, equal
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or greater number of contributors were detected in the remaining exhibit samples. In general,
exhibits with higher DNA source resulted in higher pick-up of DNA by the glove during examination. For each exhibit, except one, at least one of the gloves had higher number of
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contributors than evidentiary profiles.
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Staff or examiners were not excluded from 14% of all the gloves used during examination
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(likelihood ratio of greater than or equal to 1000 without theta was chosen as the arbitrary threshold). Of these gloves, 81% were attributed to staff directly involved in the examination
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and all were either first or last gloves. While 19% of these gloves were attributed to staff that
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were not directly involved in the exhibit examination.
Three of the four gloves with apparent non-examiner staff member non-exclusion were from
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the middle of two different examinations. The apparent non-examiner staff member was a different individual in each instance. However, the likelihood ratio in each of these instances was below the laboratory threshold of 10,000 (without theta), indicating the matches are
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considered more likely to be adventitious than a contamination event.
3.3
Control profiles
Of the 49 controls analysed, 96% did not produced a profile. Of the 4% (i.e. 2 samples) that did, one was a low level, partial single source DNA profile and one was a partial, mixed 9
DNA profile. These two profiles did not match any staff on the elimination database, POI in the case or the evidentiary profiles. This may be due to presence of low levels of DNA introduced during manufacturing [34] or handling and storage prior to use.
3.4
Video recordings
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Due to technical issues, 2 of 11 examination video recordings (both involving the same
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examiner) were not able to be analysed and thus not included in the summary of this data.
The video recordings showed that the number of contacts made by one gloved hand ranged from 1 to 52 contacts, with an average of 232 total contacts made by examiners per
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examination (includes both gloves). On average, examiners made 4.15 and 4.13 contacts per
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minute by the left and right hands respectively with various objects used throughout each
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examination. However, one examiner made significantly more contacts than the other two
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examiners.
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An exhibit was contacted, accumulatively by all gloves during an examination, on average 14.5 and 15 times by the left and right hands respectively. However, the number of contacts
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varied considerably between examiners and exhibit types with a lower range of 5 contacts, to
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an upper range of 61 contacts made with an exhibit (in total with both hands). Each left and right gloves touched on average 6.58 and 6.92 different types of objects respectively with no significant difference in contacts between left and right hands. Note, none of the examiners
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touched their skin during any of the examinations observed.
The total duration of an examination of an item that required screening for blood was significantly longer than items that required trace sampling. Furthermore, the total number of contacts made by examiners when screening for blood on an item was significantly more than items that required trace sampling. 10
The video recordings were also assessed to determine if any glove contacts could potentially pose an increased DNA contamination risk based upon examiner behaviour, including what was touched and in which sequence. The following contacts were not considered to be a risk: (a) contacts of clean glove (i.e. new gloves just put on and those that have only touched “clean” DNA-free tools) with clean tools (b) contacts of “dirty” gloves with “dirty” tools (c)
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retouching the same area with the same glove multiple times directly after each other without
touching anything in-between (although acknowledge that this could lead to lose of DNA).
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However, other types of contact were considered an increased risk and were defined into two
categories: high and low risk. High risk was defined as (a) any direct or primary contact that
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is likely to result in transfer (e.g. contacting the exhibit with gloves previously used to touch the outer packaging of the exhibit); (b) any contacts that can result in case-to-case transfer
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(e.g. contacts that could have introduced DNA into a “clean” (DNA-free) environment, such
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as touching DNA-free tools within a designated “clean” draw of tools). All other contacts were considered to be low risk (e.g. touching the description paper with the gloves previously
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used to touch the item). Approximately 7.7% of all contacts were considered to pose a potential increased contamination risk, with approximately 62% of these considered to be in
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the low risk category and 37% in the high risk category. Of the high risk category, it was noted that these contacts mainly occurred in the process of removing the exhibit from the
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packaging, or in re-packaging the exhibit.
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3.5 Casework impact assessment
An assessment of the profiles generated from the exhibits examined as part of this study and the interpretation/conclusions drawn from these as part of the associated criminal investigations did not reveal any negative impacts. Specifically, DNA profiles obtained on the gloves were compared to the relevant evidentiary DNA profiles to investigate if any
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detrimental effects, such as loss of relevant DNA to the gloves, could be noted. Further, all evidentiary profiles were investigated for the presence of staff DNA contamination, from the gloves, and none had detectable staff profiles.
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4. Discussion
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Due to the targeted sampling of the evidentiary exhibits based on case work information,
POI in the case were not excluded from the majority of the evidentiary profiles. Not
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surprisingly, these profiles were also detected on the gloves used during examination. Of
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note, POI were detected on approximately twice as many gloves used to examine blood than
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trace exhibits. Interestingly, on one occasion a partial profile of the POI was detected on the
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glove (albeit with low support) but not on the exhibit. This is most likely due to the glove touching parts of the exhibit different to the part that was sampled. Generally, a targeted
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sampling approach is taken during examination based either on information provided by the investigator, or on the knowledge of the exhibits and how they are commonly used or
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touched. However, there may be instances when target area will not correspond to the best
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region for testing. This also highlights the possibility that the absence of the DNA profile of the POI on the exhibit in question may be a result of poor identification of area to be targeted, emphasising the importance of sourcing as much case relevant information as
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possible prior to sampling (unless the entire exhibit is sampled), and/or need for improved availability / application of visualisation methodologies.
Unknown DNA alleles not attributed to persons related to the case or the examiners and staff were detected on the gloves. While some of these alleles were detected in evidentiary
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samples, the majority of these were only detected on the gloves. These alleles were most likely picked up from the exhibits but from areas that were not sampled by the examiners. The review of the recordings showed that presumably clean tools constituted the majority of contacts, suggesting that the unknown contributors on the gloves have originated from the evidentiary exhibits. Therefore, this DNA can potentially be re-distributed from the original
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area on the exhibit to other areas during examination via the gloves. Consequently, to avoid unnecessary introduction and loss of DNA from the target area of interest, we suggest that
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examiners should assess the exhibit in its entirety and make preliminary determination of
which areas will be sampled at the beginning of examination, where possible. Once such
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determination is made, examiners should avoid contacting the target sample area(s) for the
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duration of the examination other than during sample collection if possible.
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In 14% of the gloves, staff were not excluded as contributors to the profiles generated. However, this was largely contained to the first and last pairs of gloves, which also had
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more DNA and higher number of contributors. The gloves that touched the packaging were in most instances not observed subsequently touching an exhibit or clean tools, as they were
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changed prior to such actions. A possible explanation could be that DNA was transferred to the gloves as they were put on. However, because examiner’s DNA was not detected on the
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other gloves used during examination, this is unlikely. It is more likely that the DNA of the examiner was present on the packaging and then transferred to the gloves. The outer exhibit
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packaging likely contains DNA of the examiners deposited whilst collecting, carrying and storing the packaged exhibit as gloves are not worn during this period. This could also be a further source of unknown DNA as others, not on the staff elimination database may also have handled the packaging of the exhibit. The DNA on the packaging can be picked up and transferred to the gloves that come in contact with the packaging. Therefore, care must be
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taken not to touch the exhibit with the gloves that contacted the packaging in order to avoid such contamination.
Future studies that include samples from outer packaging may illuminate this level of risk. Inclusion of staff, other than the examiners, that handle packaging of exhibits, or work in
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areas where packaging and exhibits are stored, would also be of assistance to determine root
causes of apparent contamination events [see also 43,44,51]. The review of the recordings
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provides an insight into examiner behaviour; the number of contacts with the exhibit and the variation between examiners in tools used influenced by the exhibit and biological source
type. One of the high risk behaviours noted was contacts made by “dirty” gloves with tools
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and surfaces deemed to be “clean”. Such contacts may leave behind evidentiary and other
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DNA that can be picked up by subsequent examiners in future unrelated cases. However, in our laboratory all tools are cleaned just prior to commencement of examinations, even
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though they are taken from a designated “clean” area, as per internal procedure. It was also
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noted that examination gloves, on occasion, made contact with outer layer of the exhibits packaging and then touched the exhibit in the process of removing the exhibit from the
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packaging. Although DNA from the examiner was detected on the gloves used to touch the packaging, it was not detected on gloves that did not touch the packaging, or in evidentiary
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samples.
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Not only did gloves on occasion come into contact with exhibits after contacting the packaging when removing the item from the packaging, but similarly, re-packaging the exhibit appeared in some instances to cause inadvertent contact between the outer packaging and the exhibit. This could become an issue if the exhibit was later required to be re-examined. This risk can be somewhat mitigated by having clear procedures and training
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around when examiners should change gloves. Further, packaging that is easier to open, and/or able to be cleaned, may help reduce the risk of inadvertent contact in the process of removing and re-packaging the exhibit.
The approximate four contacts made by each gloved hand per minute were fewer than the
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number of contacts made by the dominant hand during everyday activities [59] and either hand during unarmed robberies [60]. This may in part be due to attended behaviours of the
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examiners associated with the task being performed. It could also be argued that examiner behaviour may have been influences by the knowledge that they were being recorded.
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However we believe that if so, such influence would tend to improve examiner behaviour
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from their normal modus operandi.
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The results of this study indicate that gloves used during examination can collect DNA from the exhibits which could prove problematic in some circumstances. For instance, during
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trace sampling, such loses to the gloves can result in the reduction of DNA available therefore impacting the quality of the evidentiary profile. Furthermore, DNA collected on
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the gloves could be redeposited on other parts of the exhibit which may be an issue during
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targeted sampling.
The examiners in this study frequently replaced their gloves at various stages during the
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examination, with new gloves. The employment of double gloving procedures (i.e. examiner wears two gloves per hand and only replaces the outer glove when deemed dirty prior to high risk touch) may be an additional means of reducing the contamination risk as the singularly gloved hands rather than naked hands are involved with putting on the outer
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second glove. The relative merits and practicalities of these methods warrant further investigation.
As previously reported [42] there is a need for processes to detect and report contamination events and to conduct root-cause analyses to help reduce future risks. It is recognised that
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different jurisdictions, crime scene attending agencies and forensic laboratories operate
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according to different contamination minimisation, and monitoring, protocols.
There is a necessity for further evaluation of exhibit examination practises including the
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timing and frequency of glove changes, the number of contacts made with the exhibit and
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the areas on the exhibits being contacted. Additionally, consideration should be given to the
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potential for DNA transfer from the outside of the packaging, which is likely to contain
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DNA from multiple sources, to the gloves and consequently to the exhibit. One possible means of reducing the uncertainty of potential loss and/or transfer of DNA via gloves
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detrimental to an investigation, could be through the collection and packaging of all gloves (separately) used during exhibit examination (thus available for profiling if deemed
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relevant) until such time as the case is closed and court proceedings are finalised. However, the feasibility of such endeavour would be logistically time consuming and costly, and the
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gloves likely to rarely be sampled, so a thorough cost benefit analyses would be recommended as part of any consideration of pursuing such a protocol. Similarly, video
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recording of examinations and DNA analysis may benefit root cause analyses where contamination is suspected. This too would require a cost benefit analyses as part of any further consideration.
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The considerations of this study should extend outside of the forensic biology laboratory and include all scientific and other personnel that come in contact with items that downstream require DNA examination. Given that ‘risky’ contacts were made by highly trained forensic biology specialists in an optimal laboratory environment, it could be extrapolated that initial collection of items by personnel who may have less
training
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regarding contamination minimisation, work within a more challenging environments, and
have fewer resources to safeguard against contamination, could result in additional risky
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contacts. Anecdotally, and supported by Fonnelop et al [51] and Hauhart and Mmenius [61], the level of caution taken and the frequency of glove changes is less for many non-
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biology/DNA practitioners who handle exhibits requiring biological examination. This is
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partly due to less awareness training and competency assessments regarding DNA transfer,
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contamination risks and contamination risk minimisation, and in some instances lack of
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appropriate formal procedures and/or equipment/tools, and/or constrained work environment. This highlights the need for further investment into on-going training and
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support (e.g. easy use/fit for purpose crime scene kits) particularly for non-crime scene/forensic scientist such as operational Police Officers. Guidelines and compliance
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requirements like those implemented by the United Kingdom Forensic Science Regulator, [62] are a useful step toward achieving desired improvement in limiting contamination risks
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and the negative consequences of contamination events.
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Acknowledgements
We would like to thank Dr Bianca Szkuta for her assistance in preparing Fig. 1.
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K. Steensma, R. Ansell, L. Clarisse, E. Connolly, A.D. Kloosterman, L.G. McKenna, R.A.H. van Oorschot, B. Szkuta, B. Kokshoorn, An inter-laboratory comparison study on transfer, persistence and recovery of DNA from cable ties, Forensic Science International: Genetics, 31 (2017) 95-104.
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R.K. Farmen, R. Jagho, P. Cortez, E.S. Froyland, Assessment of individual shedder status and implications for secondary DNA transfer, Forensic Science International: Genetics Supplement Series 1 (1) (2008) 415-417.
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R. Daniel, R.A.H. van Oorschot, An investigation of the presence of DNA on unused laboratory gloves, Forensic Science International: Genetics Supplement Series 3 (2011) e45-e46.
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K.N. Ballantyne, R. Salemi, F. Guarino, J.R. Pearson, D. Garlepp, S Fowler, R.A.H. van Oorschot, DNA contamination minimisation - finding an effective cleaning method. Aust. J. Forensic Sci. 47(4) (2015) 428-439.
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Figures
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Fig. 1. A diagrammatic representation of all the contacts made by all the gloves worn during an examination of one item and the DNA results obtained from them. All times are approximate and rounded to the nearest minute. The total number of alleles detected on each glove, excluding artefacts, represented in brackets. (Note: the duration of each touch, where the object(s) was contacted and with which part of the glove, are not included here). Unless indicated (staff or POI) the DNA detected on the gloves was from an unknown source.
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Glove symbol: indicates a change of glove
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= contact with possible risk P = number of people contributing to the DNA profile; e.g. 3P= three person mixture (with the total number of alleles detected represented in brackets)
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Fig.2. Parts of DNA profiles generated from three gloves: (A) the glove used in trace exhibit examination from which a POI in the case was not excluded with the LR of 22* (POI excluded from the evidentiary profile); (B) the last glove used in trace exhibit examination that was used to re-package the exhibit from which the examiner* was not excluded; (C) the first glove used in trace exhibit examination that was used to touch the packaging and objects utilized for the examination (high risk contacts) from which the POI and staff were excluded (ci) and a corresponding part of a DNA evidence profile generated from the exhibit (cii)
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Note: only part of the profile was used and loci and allele designations were masked for confidentiality purposes
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* POI and examiner alleles (as applicable) are identified by blue boxes with black boxes used to mask allele designation where all DNA reference samples were excluded
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No. of gloves used examining trace exhibits (pairs of gloves) 98 (49 pairs)
No. of gloves with alleles above threshold 170
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193 (96 pairs)a
No. of gloves used examining blood exhibits (pair of gloves) 95 (46 pairs)a
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No. of gloves worn and sampled (pairs of gloves)
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Table 1 Summary of number of gloves used and the number of gloves providing specific types of DNA result No. of gloves with alleles above threshold with unknownb DNA 131
In one instance a single glove was changed rather than both gloves of a pair
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Unknown= alleles not matching POI’s in the case or staff and examiner
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a
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Table 2. Summary of experiment data relating to the duration of examination, number of contacts by gloves and the DNA quantity and profiles, per examiner and exhibit
a
34 48 66 51 83 54 52 140 N/A 68 N/A
No. of contacts Total
No. of Exhibit contacts
177 158 232 205 305 308 304 255 N/A 144 N/A
21 5 45 45 52 61 18 30 N/A 18 N/A
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A B B A A A A C D C D
No. of Pairs of gloves (n)
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Exam. Duration (min)
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Examiner
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1 2 3 4 5 6 7 8 9 10 11
Exhibit type Trace (T) or Blood (B) T T T B B B B T B T B
PT
Item
4 8 14 8 7 6 5 15 9.5a 8 12
Total DNA quant Average (ng) 0.177 0.007 0.009 0.361 0.489 0.261 0.138 0.126 0.357 0.098 0.245
Nine pairs and one single glove
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N/A = examinations where video recording failed
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Total DNA quant Range (ng) 0-0.53 0-0.310 0-0.118 0-1.499 0-0.132 0-1.511 0-0.459 0-0.42 0-1.02 0-0.9 0-1.08
% gloves with alleles above detection threshold 100 81 39 100 100 100 100 90 95 100 100
No. of alleles Averag e
No. of allele Range
No. of contributors Average
No of contributors Range
27 12 2 48 49 28 26 22 42 15 35
1-101 0-42 0-18 2-107 2-121 1-98 6-64 0-64 2-73 1-64 1-76
2 1 0.5 3 4 2 2 2 2 2 2
1-5 0-3 0-2 1-5 1-6 1-4 1-4 0-3 0-4 1-3 1-4
S1872-4973(18)30428-9 https://doi.org/10.1016/j.fsigen.2018.10.018 FSIGEN 1990
To appear in:
Forensic Science International: Genetics
Received date: Revised date: Accepted date:
31 July 2018 26 October 2018 31 October 2018
Please cite this article as: Goray M, Pirie E, van Oorschot RAH, DNA transfer: DNA acquired by gloves during casework examinations, Forensic Science International: Genetics (2018), https://doi.org/10.1016/j.fsigen.2018.10.018 This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
DNA transfer: DNA acquired by gloves during casework examinations Mariya Goraya,*, Erin Piriea and Roland A.H. van Oorschotb,c
Biometric Division, Victoria Police Forensic Services Department, Macleod, Australia
b
Office of the Chief Forensic Scientist, Victoria Police Forensic Services Department,
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a
c
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Macleod, Australia
School of Molecular Sciences, College of Science, Health and Engineering, La Trobe
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University, Bundoora, Australia
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*Corresponding author: Telephone: +61394509715
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Email: [email protected]
Highlights
Examiner variation in exhibit areas touched and timing of glove changes
Exhibit and staff derived DNA acquired by gloves during examinations
Risk of removal, addition and redistribution of DNA by gloves during examination
Examiners need to be cognisant of areas touched and timing of glove changes
Stages of removing exhibit from packaging and repackaging require special attention
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1. Introduction
DNA transfer prior, during and post criminal activity has been a topic of conversation among
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forensic scientists for some time now [1-38]. Questions relating to when and how the DNA material was deposited onto surfaces of interest are becoming paramount [39-41]. These
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questions are pertinent to the events relating to criminal activity as well as the activities of
investigators post criminal event. Contamination post criminal event by investigators have
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been observed [42-44] and can occur through various means, including: non- or inappropriate
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use of personal protective equipment when near and/or handling exhibits to be examined;
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contact of such exhibits with dirty tools and/or equipment [2, 45-52]. Here we focus on
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elements relating to the potential of DNA transfer via gloves during examination of exhibits
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by examiners within a forensic biology laboratory.
Most surfaces and objects that are regularly contacted will have some level of background
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DNA [22, 53-54]. The quality and quantity of this DNA will depend on the frequency with
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which these surfaces are touched/contacted, by whom/what and the type and regularity of cleaning involved [48, 52, 54-56]. Environmental DNA studies of forensic biology laboratories have demonstrated that surface, equipment and tools within the laboratory can
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also contain detectable quantities of DNA, that may have been derived from examiners, others that have entered the area or handled the packaging, equipment, tools, consumables, or from exhibits previously examined [48,52,52,54]. Unclean surfaces, equipment and tools within a forensic laboratory pose a potential risk in respect to contamination of exhibits to be examined.
2
Contamination of exhibits can occur at any stage of an investigation. Gloves worn during examination are a key risk factor for such contamination. This risk will be dependent on how gloves are put on, what they touch and when they are replaced. Addition, loss and/or redistribution of DNA to/from/within an exhibit through handling during examination can
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have significant implications for the investigation, placing doubt on the meaning of any
results obtained [15, 37-48, 50-52]. Several studies have investigated potential DNA transfer
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during mock “casework” examination (for example with tools), allowing for some inferences of what can be expected in real casework. However, to our knowledge, no studies have
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investigated DNA transfer via gloves during actual casework examination.
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The aim of this study is to investigate the risk of DNA loss, addition and redistribution via
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gloves during real casework examination, as well as the appropriateness and frequency of glove changes by the examiners. Several examiners were video recorded during examination
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of a range of casework exhibits. The timing of all glove changes and all contacts made by each glove were documented. Each glove used during the examination was sampled at the
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moment of the intended glove removal. The profiles generated from these gloves were examined in combination with the profiles generated from the exhibits, persons relevant to
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the case, the examiner, the researchers and other staff.
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2. Materials and Methods
2.1
Experimental set up
New gloves (“InControl” long cuff nitrile (power free) gloves) were sampled (as negative controls) and placed in dedicated glove boxes with each six gloves separated by blotting
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paper, ready for use. The glove box openings were widened in order to minimize examiner contacts with the box and the gloves. The boxes were placed into new clip seal bags to minimize exposure until required. The unused gloves remaining within a blotting paper section were discarded after each examination. Four examiners (A-D) were each videorecorded during exhibit examination and each pair of gloves used by the examiners during
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examination starting with the pair used to open the exhibit packaging and finishing with the pair used to return the exhibit back to its packaging were separately sampled (left and right).
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The palm and fingers (palm side only and avoiding the cuff area) of each glove were sampled
by a researcher whilst still being worn by the examiner and the samples then processed as per
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case work protocols.
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Eleven separate examinations were conducted (5 by examiner A, 2 by examiner B, 2 by
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examiner C and 2 by examiner D). Each examination was of a single exhibit. Of these
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exhibits, 5 were examined for trace (metal pole, section of mattress, jeans, jacket, and bottle opener) and 6 for blood (jacket, hockey stick, two singlets, knife and t-shirt). Overall, 96
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pairs of gloves from the examination of 11 exhibits were analysed.
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The number, duration, sequence and types of contracts made by each glove during
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examination were documented from the video recordings and assessed against the DNA profiles obtained from the gloves as well as those obtained from samples collected from the
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exhibits by the examiners.
2.2
Sample Processing
Each glove was swabbed separately (wet and dry swabs) (150C swabs, Copan, USA) and the two swabs from each glove were placed directly into a 2ml tube for further processing. DNA was extracted, quantified and amplified as per manufacturer recommendations, using DNA
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IQTM (Promega), Quantifiler®Trio (Applied Biosystems) and PowerPlex® 21 (Promega) kits, respectively. Amplified products were run on a 3500xL Genetic Analyser (Applied Biosystems) and typed using GeneMapperTM IDx Software (Applied Biosystems) with a baseline threshold of 175 RFU.
Data Interpretation and Statistical Analysis
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2.3
The total DNA retrieved from each glove was calculated by multiplying the DNA
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concentration, as determined during quantification, by the volume of the extract (60 µl). The total number of alleles for each sample was determined by counting the total number of
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alleles detected at each locus (threshold: 175 RFU), excluding peaks that were determined to
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be potential artefacts (e.g. stutter) as per the laboratory interpretation method and alleles at
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Amelogenin locus.
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The minimum number of contributors was determined for each of the profiles obtained, as per our laboratory’s casework DNA interpretation method, using a modified allele count
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method which takes into consideration stochastic effects. Profiles were then interpreted and
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statistically evaluated using continuous probabilistic software STRmixTM (version 2.06) [5758]. This software weights genotype combinations and allows comparison to persons of
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interest (POI) and the staff elimination database, expressed as likelihood ratios. Furthermore, STRmixTM interpretation output provides additional information such as; the mixture proportions expressed as percentages. The comparison to the staff elimination database via
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STRmix involves calculating a Likelihood Ratio to each person on the database; without the use of theta.
The Kruskal-Wallis one-way analysis of variance was used to test if the k independent samples were from the same or different populations (at p<0.05 significance level). Given
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the continuous nature of the variables examined, statistically significant groups were analysed with the Mann-Whitney U-test and the relationship between the continuous variables using Spearman’s correlation (at p<0.05 significance level). Pearson’s correlation was used to measure the strength and direction of association that exists between two variables. Statistical
2.4
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analyses were performed in SPSS Statistics (IBM).
Quality Controls and Cleaning Protocols
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The laboratory space and tools were cleaned as per laboratory protocols with 1%
Hypochlorite followed by distilled water. Of all the control swabs taken from gloves used by
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the examiners prior to them being worn, forty-nine (chosen based on the DNA result from the
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gloves) were processed to check for possible contamination. Twenty-four control swabs from
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the gloves where staff were not excluded as contributors to the DNA obtained were chosen
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for analysis. The remaining twenty-five controls processed were chosen randomly. Both positive and negative controls were implemented during DNA processing steps as per
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3. Results
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casework methods and produced expected results.
3.1
General
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Figure 1 is an illustrative snapshot of some of the data collected during one of the observed examinations.
Ninety-five gloves were used to examine six blood exhibits and ninety-eight gloves to examine five trace exhibits. The number of glove changes per exhibit ranged from 4 to 15
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pairs (average 9). On average 10 pairs of gloves were used during blood exhibits examination and 8 during trace exhibit examinations. Total DNA detected on the gloves ranged from 0 to 2.26ng (average 0.21ng) with an average of 0.321ng detected on the blood exhibit gloves and 0.106ng detected on the trace exhibit gloves. Profiles were produced from 98% of blood exhibit gloves and 87% of trace exhibit gloves. The remaining 2% and 23% respectively
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(total 25 gloves) did not contain sufficient DNA to produce a DNA profile. The total number of alleles ranged from 0 to 121 (average 26). There was a statistically significant positive
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correlation between the number of alleles and total DNA detected. The number of contributors on the gloves ranged from 0 to 6 people (average 1.8). The average number of
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contributors on blood and trace exhibit gloves was 2.3 and 1.3, respectively. In general, the
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gloves used at the beginning and those used at the end of an examination had approximately
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twice as much DNA and number of contributors than other gloves.
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The total quantity of DNA, total number of alleles, and the number of contributors detected on the gloves used to examine blood exhibits were significantly greater than on the gloves
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used to examine trace exhibits. There were no significant differences in the total quantity of
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DNA, the total number of alleles, and the number of contributors detected between the left
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and right hand gloves.
When comparing the glove changes between examiners, three examiners had an average of 11 glove changes for blood and trace exhibits combined, and one examiner had an average of
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6 changes per examination. There were significant differences in total DNA quantities, total number of alleles and number of contributors detected on the gloves among the four examiners. Two examiners consistently had significantly more DNA, more alleles and higher number of contributors detected on the gloves when compared to the other two examiners.
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On average, there were six glove changes for soft/flexible exhibits including clothing and piece of mattress and ten glove changes for hard/solid exhibits including weapons, metal pole etc. There were no significant differences in total DNA quantities, total number of alleles and number of contributors detected on the gloves between the soft/flexible and hard/solid
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exhibits.
Of the gloves with alleles above the detection threshold 78% (131 of 170 gloves) had at least
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one unknown allele detected that could not be attributed to DNA reference samples relevant
to the case, the evidentiary profiles obtained from the exhibit or to the examiner. Sixty five percent of the gloves with unknown DNA alleles were from gloves used to examine blood
Comparisons with evidentiary, reference, staff and examiner profiles
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3.2
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exhibits.
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The profiles generated from the gloves were compared to the profiles obtained from the evidentiary exhibits, the DNA reference samples relevant to each exhibit and case as well as
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profiles of the examiners and those on the staff elimination database.
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In 54% and 27% of blood and trace exhibit gloves respectively the POI relevant to the case
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(either suspects and/or complainants) was not excluded as contributor(s) to the DNA results obtained. Complainant was POI on weapons and suspect’s clothing; while suspect was POI on the complainant’s clothing and weapons. No POI DNA was detected on two (metal pole,
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section of mattress) of the five trace exhibits; for the remaining three exhibits (jeans, jacket, bottle opener), the POI was not excluded in 1/28, 18/30 and 3/16 gloves respectively. In most instances where the POI was not excluded from the gloves, they were also not excluded from the evidentiary profiles generated during exhibit sampling. Interestingly, for one of the
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exhibits, a bottle opener, the suspect was not excluded, albeit with low support, (LR=22, with theta) from the gloves, yet was excluded from the evidentiary profile.
Comparison of the results generated from the gloves and exhibits showed that in most instances higher total amounts of DNA were detected in the evidentiary samples compared to
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the examination gloves (for both blood and trace gloves). Further, while 33% of the gloves had higher number of contributors compared to the corresponding evidentiary profiles, equal
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or greater number of contributors were detected in the remaining exhibit samples. In general,
exhibits with higher DNA source resulted in higher pick-up of DNA by the glove during examination. For each exhibit, except one, at least one of the gloves had higher number of
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contributors than evidentiary profiles.
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Staff or examiners were not excluded from 14% of all the gloves used during examination
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(likelihood ratio of greater than or equal to 1000 without theta was chosen as the arbitrary threshold). Of these gloves, 81% were attributed to staff directly involved in the examination
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and all were either first or last gloves. While 19% of these gloves were attributed to staff that
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were not directly involved in the exhibit examination.
Three of the four gloves with apparent non-examiner staff member non-exclusion were from
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the middle of two different examinations. The apparent non-examiner staff member was a different individual in each instance. However, the likelihood ratio in each of these instances was below the laboratory threshold of 10,000 (without theta), indicating the matches are
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considered more likely to be adventitious than a contamination event.
3.3
Control profiles
Of the 49 controls analysed, 96% did not produced a profile. Of the 4% (i.e. 2 samples) that did, one was a low level, partial single source DNA profile and one was a partial, mixed 9
DNA profile. These two profiles did not match any staff on the elimination database, POI in the case or the evidentiary profiles. This may be due to presence of low levels of DNA introduced during manufacturing [34] or handling and storage prior to use.
3.4
Video recordings
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Due to technical issues, 2 of 11 examination video recordings (both involving the same
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examiner) were not able to be analysed and thus not included in the summary of this data.
The video recordings showed that the number of contacts made by one gloved hand ranged from 1 to 52 contacts, with an average of 232 total contacts made by examiners per
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examination (includes both gloves). On average, examiners made 4.15 and 4.13 contacts per
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minute by the left and right hands respectively with various objects used throughout each
A
examination. However, one examiner made significantly more contacts than the other two
M
examiners.
ED
An exhibit was contacted, accumulatively by all gloves during an examination, on average 14.5 and 15 times by the left and right hands respectively. However, the number of contacts
PT
varied considerably between examiners and exhibit types with a lower range of 5 contacts, to
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an upper range of 61 contacts made with an exhibit (in total with both hands). Each left and right gloves touched on average 6.58 and 6.92 different types of objects respectively with no significant difference in contacts between left and right hands. Note, none of the examiners
A
touched their skin during any of the examinations observed.
The total duration of an examination of an item that required screening for blood was significantly longer than items that required trace sampling. Furthermore, the total number of contacts made by examiners when screening for blood on an item was significantly more than items that required trace sampling. 10
The video recordings were also assessed to determine if any glove contacts could potentially pose an increased DNA contamination risk based upon examiner behaviour, including what was touched and in which sequence. The following contacts were not considered to be a risk: (a) contacts of clean glove (i.e. new gloves just put on and those that have only touched “clean” DNA-free tools) with clean tools (b) contacts of “dirty” gloves with “dirty” tools (c)
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retouching the same area with the same glove multiple times directly after each other without
touching anything in-between (although acknowledge that this could lead to lose of DNA).
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However, other types of contact were considered an increased risk and were defined into two
categories: high and low risk. High risk was defined as (a) any direct or primary contact that
U
is likely to result in transfer (e.g. contacting the exhibit with gloves previously used to touch the outer packaging of the exhibit); (b) any contacts that can result in case-to-case transfer
N
(e.g. contacts that could have introduced DNA into a “clean” (DNA-free) environment, such
M
A
as touching DNA-free tools within a designated “clean” draw of tools). All other contacts were considered to be low risk (e.g. touching the description paper with the gloves previously
ED
used to touch the item). Approximately 7.7% of all contacts were considered to pose a potential increased contamination risk, with approximately 62% of these considered to be in
PT
the low risk category and 37% in the high risk category. Of the high risk category, it was noted that these contacts mainly occurred in the process of removing the exhibit from the
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packaging, or in re-packaging the exhibit.
A
3.5 Casework impact assessment
An assessment of the profiles generated from the exhibits examined as part of this study and the interpretation/conclusions drawn from these as part of the associated criminal investigations did not reveal any negative impacts. Specifically, DNA profiles obtained on the gloves were compared to the relevant evidentiary DNA profiles to investigate if any
11
detrimental effects, such as loss of relevant DNA to the gloves, could be noted. Further, all evidentiary profiles were investigated for the presence of staff DNA contamination, from the gloves, and none had detectable staff profiles.
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4. Discussion
SC R
Due to the targeted sampling of the evidentiary exhibits based on case work information,
POI in the case were not excluded from the majority of the evidentiary profiles. Not
U
surprisingly, these profiles were also detected on the gloves used during examination. Of
N
note, POI were detected on approximately twice as many gloves used to examine blood than
A
trace exhibits. Interestingly, on one occasion a partial profile of the POI was detected on the
M
glove (albeit with low support) but not on the exhibit. This is most likely due to the glove touching parts of the exhibit different to the part that was sampled. Generally, a targeted
ED
sampling approach is taken during examination based either on information provided by the investigator, or on the knowledge of the exhibits and how they are commonly used or
PT
touched. However, there may be instances when target area will not correspond to the best
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region for testing. This also highlights the possibility that the absence of the DNA profile of the POI on the exhibit in question may be a result of poor identification of area to be targeted, emphasising the importance of sourcing as much case relevant information as
A
possible prior to sampling (unless the entire exhibit is sampled), and/or need for improved availability / application of visualisation methodologies.
Unknown DNA alleles not attributed to persons related to the case or the examiners and staff were detected on the gloves. While some of these alleles were detected in evidentiary
12
samples, the majority of these were only detected on the gloves. These alleles were most likely picked up from the exhibits but from areas that were not sampled by the examiners. The review of the recordings showed that presumably clean tools constituted the majority of contacts, suggesting that the unknown contributors on the gloves have originated from the evidentiary exhibits. Therefore, this DNA can potentially be re-distributed from the original
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area on the exhibit to other areas during examination via the gloves. Consequently, to avoid unnecessary introduction and loss of DNA from the target area of interest, we suggest that
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examiners should assess the exhibit in its entirety and make preliminary determination of
which areas will be sampled at the beginning of examination, where possible. Once such
U
determination is made, examiners should avoid contacting the target sample area(s) for the
A
N
duration of the examination other than during sample collection if possible.
M
In 14% of the gloves, staff were not excluded as contributors to the profiles generated. However, this was largely contained to the first and last pairs of gloves, which also had
ED
more DNA and higher number of contributors. The gloves that touched the packaging were in most instances not observed subsequently touching an exhibit or clean tools, as they were
PT
changed prior to such actions. A possible explanation could be that DNA was transferred to the gloves as they were put on. However, because examiner’s DNA was not detected on the
CC E
other gloves used during examination, this is unlikely. It is more likely that the DNA of the examiner was present on the packaging and then transferred to the gloves. The outer exhibit
A
packaging likely contains DNA of the examiners deposited whilst collecting, carrying and storing the packaged exhibit as gloves are not worn during this period. This could also be a further source of unknown DNA as others, not on the staff elimination database may also have handled the packaging of the exhibit. The DNA on the packaging can be picked up and transferred to the gloves that come in contact with the packaging. Therefore, care must be
13
taken not to touch the exhibit with the gloves that contacted the packaging in order to avoid such contamination.
Future studies that include samples from outer packaging may illuminate this level of risk. Inclusion of staff, other than the examiners, that handle packaging of exhibits, or work in
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areas where packaging and exhibits are stored, would also be of assistance to determine root
causes of apparent contamination events [see also 43,44,51]. The review of the recordings
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provides an insight into examiner behaviour; the number of contacts with the exhibit and the variation between examiners in tools used influenced by the exhibit and biological source
type. One of the high risk behaviours noted was contacts made by “dirty” gloves with tools
N
U
and surfaces deemed to be “clean”. Such contacts may leave behind evidentiary and other
A
DNA that can be picked up by subsequent examiners in future unrelated cases. However, in our laboratory all tools are cleaned just prior to commencement of examinations, even
M
though they are taken from a designated “clean” area, as per internal procedure. It was also
ED
noted that examination gloves, on occasion, made contact with outer layer of the exhibits packaging and then touched the exhibit in the process of removing the exhibit from the
PT
packaging. Although DNA from the examiner was detected on the gloves used to touch the packaging, it was not detected on gloves that did not touch the packaging, or in evidentiary
CC E
samples.
A
Not only did gloves on occasion come into contact with exhibits after contacting the packaging when removing the item from the packaging, but similarly, re-packaging the exhibit appeared in some instances to cause inadvertent contact between the outer packaging and the exhibit. This could become an issue if the exhibit was later required to be re-examined. This risk can be somewhat mitigated by having clear procedures and training
14
around when examiners should change gloves. Further, packaging that is easier to open, and/or able to be cleaned, may help reduce the risk of inadvertent contact in the process of removing and re-packaging the exhibit.
The approximate four contacts made by each gloved hand per minute were fewer than the
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number of contacts made by the dominant hand during everyday activities [59] and either hand during unarmed robberies [60]. This may in part be due to attended behaviours of the
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examiners associated with the task being performed. It could also be argued that examiner behaviour may have been influences by the knowledge that they were being recorded.
U
However we believe that if so, such influence would tend to improve examiner behaviour
A
N
from their normal modus operandi.
M
The results of this study indicate that gloves used during examination can collect DNA from the exhibits which could prove problematic in some circumstances. For instance, during
ED
trace sampling, such loses to the gloves can result in the reduction of DNA available therefore impacting the quality of the evidentiary profile. Furthermore, DNA collected on
PT
the gloves could be redeposited on other parts of the exhibit which may be an issue during
CC E
targeted sampling.
The examiners in this study frequently replaced their gloves at various stages during the
A
examination, with new gloves. The employment of double gloving procedures (i.e. examiner wears two gloves per hand and only replaces the outer glove when deemed dirty prior to high risk touch) may be an additional means of reducing the contamination risk as the singularly gloved hands rather than naked hands are involved with putting on the outer
15
second glove. The relative merits and practicalities of these methods warrant further investigation.
As previously reported [42] there is a need for processes to detect and report contamination events and to conduct root-cause analyses to help reduce future risks. It is recognised that
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different jurisdictions, crime scene attending agencies and forensic laboratories operate
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according to different contamination minimisation, and monitoring, protocols.
There is a necessity for further evaluation of exhibit examination practises including the
U
timing and frequency of glove changes, the number of contacts made with the exhibit and
N
the areas on the exhibits being contacted. Additionally, consideration should be given to the
A
potential for DNA transfer from the outside of the packaging, which is likely to contain
M
DNA from multiple sources, to the gloves and consequently to the exhibit. One possible means of reducing the uncertainty of potential loss and/or transfer of DNA via gloves
ED
detrimental to an investigation, could be through the collection and packaging of all gloves (separately) used during exhibit examination (thus available for profiling if deemed
PT
relevant) until such time as the case is closed and court proceedings are finalised. However, the feasibility of such endeavour would be logistically time consuming and costly, and the
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gloves likely to rarely be sampled, so a thorough cost benefit analyses would be recommended as part of any consideration of pursuing such a protocol. Similarly, video
A
recording of examinations and DNA analysis may benefit root cause analyses where contamination is suspected. This too would require a cost benefit analyses as part of any further consideration.
16
The considerations of this study should extend outside of the forensic biology laboratory and include all scientific and other personnel that come in contact with items that downstream require DNA examination. Given that ‘risky’ contacts were made by highly trained forensic biology specialists in an optimal laboratory environment, it could be extrapolated that initial collection of items by personnel who may have less
training
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regarding contamination minimisation, work within a more challenging environments, and
have fewer resources to safeguard against contamination, could result in additional risky
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contacts. Anecdotally, and supported by Fonnelop et al [51] and Hauhart and Mmenius [61], the level of caution taken and the frequency of glove changes is less for many non-
U
biology/DNA practitioners who handle exhibits requiring biological examination. This is
N
partly due to less awareness training and competency assessments regarding DNA transfer,
A
contamination risks and contamination risk minimisation, and in some instances lack of
M
appropriate formal procedures and/or equipment/tools, and/or constrained work environment. This highlights the need for further investment into on-going training and
ED
support (e.g. easy use/fit for purpose crime scene kits) particularly for non-crime scene/forensic scientist such as operational Police Officers. Guidelines and compliance
PT
requirements like those implemented by the United Kingdom Forensic Science Regulator, [62] are a useful step toward achieving desired improvement in limiting contamination risks
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and the negative consequences of contamination events.
A
Acknowledgements
We would like to thank Dr Bianca Szkuta for her assistance in preparing Fig. 1.
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Figures
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Fig. 1. A diagrammatic representation of all the contacts made by all the gloves worn during an examination of one item and the DNA results obtained from them. All times are approximate and rounded to the nearest minute. The total number of alleles detected on each glove, excluding artefacts, represented in brackets. (Note: the duration of each touch, where the object(s) was contacted and with which part of the glove, are not included here). Unless indicated (staff or POI) the DNA detected on the gloves was from an unknown source.
N
Glove symbol: indicates a change of glove
A
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PT
ED
M
A
= contact with possible risk P = number of people contributing to the DNA profile; e.g. 3P= three person mixture (with the total number of alleles detected represented in brackets)
23
IP T SC R U N A
PT
ED
M
Fig.2. Parts of DNA profiles generated from three gloves: (A) the glove used in trace exhibit examination from which a POI in the case was not excluded with the LR of 22* (POI excluded from the evidentiary profile); (B) the last glove used in trace exhibit examination that was used to re-package the exhibit from which the examiner* was not excluded; (C) the first glove used in trace exhibit examination that was used to touch the packaging and objects utilized for the examination (high risk contacts) from which the POI and staff were excluded (ci) and a corresponding part of a DNA evidence profile generated from the exhibit (cii)
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Note: only part of the profile was used and loci and allele designations were masked for confidentiality purposes
A
* POI and examiner alleles (as applicable) are identified by blue boxes with black boxes used to mask allele designation where all DNA reference samples were excluded
24
IP T SC R
A
No. of gloves used examining trace exhibits (pairs of gloves) 98 (49 pairs)
No. of gloves with alleles above threshold 170
ED
193 (96 pairs)a
No. of gloves used examining blood exhibits (pair of gloves) 95 (46 pairs)a
M
No. of gloves worn and sampled (pairs of gloves)
N
U
Table 1 Summary of number of gloves used and the number of gloves providing specific types of DNA result No. of gloves with alleles above threshold with unknownb DNA 131
In one instance a single glove was changed rather than both gloves of a pair
b
Unknown= alleles not matching POI’s in the case or staff and examiner
A
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PT
a
25
I N U SC R
Table 2. Summary of experiment data relating to the duration of examination, number of contacts by gloves and the DNA quantity and profiles, per examiner and exhibit
a
34 48 66 51 83 54 52 140 N/A 68 N/A
No. of contacts Total
No. of Exhibit contacts
177 158 232 205 305 308 304 255 N/A 144 N/A
21 5 45 45 52 61 18 30 N/A 18 N/A
A
A B B A A A A C D C D
No. of Pairs of gloves (n)
M
Exam. Duration (min)
ED
Examiner
CC E
1 2 3 4 5 6 7 8 9 10 11
Exhibit type Trace (T) or Blood (B) T T T B B B B T B T B
PT
Item
4 8 14 8 7 6 5 15 9.5a 8 12
Total DNA quant Average (ng) 0.177 0.007 0.009 0.361 0.489 0.261 0.138 0.126 0.357 0.098 0.245
Nine pairs and one single glove
A
N/A = examinations where video recording failed
26
Total DNA quant Range (ng) 0-0.53 0-0.310 0-0.118 0-1.499 0-0.132 0-1.511 0-0.459 0-0.42 0-1.02 0-0.9 0-1.08
% gloves with alleles above detection threshold 100 81 39 100 100 100 100 90 95 100 100
No. of alleles Averag e
No. of allele Range
No. of contributors Average
No of contributors Range
27 12 2 48 49 28 26 22 42 15 35
1-101 0-42 0-18 2-107 2-121 1-98 6-64 0-64 2-73 1-64 1-76
2 1 0.5 3 4 2 2 2 2 2 2
1-5 0-3 0-2 1-5 1-6 1-4 1-4 0-3 0-4 1-3 1-4