META-ANALYSIS OF THE SECONDARY TRANSFER OF DNA · 5 1. INTRODUCTION Deoxyribonucleic acid (DNA) is...

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META-ANALYSISOFTHESECONDARYTRANSFEROFDNA

By

TaraDUNHILL

Athesissubmittedinfulfilmentoftherequirementsforthedegreeof

MasterofForensicScience(ProfessionalPractice)

in

TheSchoolofVeterinaryandLifeSciences

MurdochUniversity

Supervisor:Mr.BrendanChapman

Semester1,2018

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DECLERATION

Ideclarethatthismanuscriptdoesnotcontainanymaterialsubmittedpreviouslyforthe

award of any other degree or diploma at any university or other tertiary institution.

Furthermore, to thebestofmyknowledge, itdoesnot containanymaterialpreviously

publishedorwrittenbyanotherindividual,exceptwhereduereferenceshasbeenmadein

thetext.Finally, Ideclarethatallreportedexperimentationsperformedinthisresearch

were carried out bymyself, except that any contribution by others,withwhom I have

workedisexplicitlyacknowledged.

Signed:TaraDunhill

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ACKNOWLEDGEMENTS

I would like to acknowledge firstlyMr. Brendan Chapman for his ongoing support and

mentorshipforthisdissertation.Secondly,Iwouldliketoacknowledgeallmyfamilyand

friends for their support and encouragement despite being over the other side of the

country,inparticularJessicaGeorgakisandAshleySymons.

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TABLEOFCONTENTS

TITLEPAGE………………………………………………………………………………………………………………………i

DECLERATION………………………………………………………………………………………………………………….ii

ACKNOWLEDGEMENTS…………………………………………………………………………………………………..iii

PARTONE

LITERATUREREVIEW…………………………………………………………………………………………………1-47

PARTTWO

MANUSCRIPT……………………………………………………………………………………………………………49-79

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THISPAGEHASBEENLEFTINTENTIONALLYBLANK

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PARTONE

LITERATUREREVIEW


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ABSTRACT

Deoxyribonucleicacid(DNA)iscomplexmoleculepresentinnearlyeverycellofthehuman

bodythatcontainsallofanindividual’sgeneticmaterial.TraceDNAhasallowedDNAtobe

detectedoneverydayobjectssuchasdoorhandles,tablesandchairs.Currently,theidea

ofthesecondarytransferofDNAhasarisen,involvingthetransferofgeneticprofilesfrom

anindividualthroughavectorandthentofinalindividualorobject.Thishasbecomeare-

occurringterminthecourtsystem,asindividualsstandingtrialusethisargumentforthe

presence of their DNA. It is therefore hypothesized that through the exploration of

literatureandcompletionofameta-analysisthatanappropriateandconciseguidewillbe

produced to determine the chances of a secondary transfer event occurring andwhat

conditions are required, to aid Biologists in expert testimony. Database searcheswere

conductedtogainresourcesinthetopic,followedbyanumberofscreenstodetermine

the suitability of articles for the meta-analysis. With the resultant 38 articles, data

extractionwasadditionallycarried.However,duetothelackofquantitativevariablesand

results,ameta-analysiswasnotabletobeconducted.It issuggestedthatinthefuture,

studiespublishtheirresultsinamorescientificallyrigorousmannerorallowtheaccessof

rawresultsforexternalinterpretationpurposes.

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TABLEOFCONTENTS

ABSTRACT…………………………………………………………………………………………………………………….2LISTOFABBREVIATIONS……………………………………………………………………………………………….41. INTRODUCTION………………………………………………………………………………………………………5

1.1 THETRANSFEROFDNA…………………………………………………………………………………..61.2 META-ANALYSIS……………………………………………………………………………………………..8

2. LITERATUREREVIEW………………………………………………………………………………...............112.1 OBJECTASVECTOR……………………………………………………………………………………….12

2.1.1 LAUNDERING……………………………………………………............................162.1.2 FINGERPRINTBRUSHES………………………………………………………….......17

2.2 INDIVIDUALASVECTOR…………………………………………………………………………........182.3 INDIRECTFINDINGSOFSECONDARYTRANSFER…………………………………………....212.4 SHEDDERSTATUS………………………………………………………………………….................252.5 OTHERFORMSOFDNAINVOLVEDINSECONDARYTRANSFER……………...........26

2.5.1 TRANSFEROFSPERMATOZA………………………………………..................262.5.2 TRANSFEROFSALIVA…………………………………………………………..........272.5.3 TRANSFEROFBLOOD……………………………………………...……...............29

2.6 TERTIARYTRANSFEROFDNAANDBEYOND……………………………….....................302.7 CASESTUDIES…………………………………………………………………………………...............32

3. PERFORMANCEOFMETA-ANALYSIS……………………………………………………..................333.1METHOD……………………………………………………………………………………....................33 3.1.1SCREENINGOFARTICLES…………………………………………….....................34 3.1.2DATAEXTRACTION………………………………………………………….................353.2RESULTSANDDISCUSSION…………………………………………………………....................36

4. LIMITATIONSOFTHESECONDARYTRANSFEROFDNA…………………………..................384.1CONTROLLEDNATUREOFTHESTUDIES………………………………….........................384.2TIMEBETWEENTAKINGSAMPLES……………………………………………......................394.3REFERENCESAMPLES………………………………………………………………......................40

5. LIMITATIONSOFMETA-ANALYSIS………………………………………………………...................405.1NEWAPPLICATIONTOTHISFIELD………………………………………………....................405.2TYPEOFRESULTSPUBLISHED………………………………………………………..................41

6. THEFUTUREFORTHESECONDARYTRANSFEROFDNA……………………......................417. CONCLUSION………………………………………………………………………………….......................418. REFERENCES…………………………………………………………………………................................43

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LISTOFABBREVIATIONS

DNA=DeoxyribonucleicAcid

ng=Nanograms

ng/µL=Nanogramspermicroliter(concentration)

pg=Pictograms

STR=Shorttandemrepeat

µL=Microliter

UV=Ultraviolet

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1. INTRODUCTION


bodythatpossessallofanindividual’sgeneticmaterial.1Eachindividualhasapproximately

threebillionbasepairswhichisessentialforthecharacterizationofeachindividual.1Of

thesethreebillionbasepairs,itisthoughtthatapproximatelyonly10%containsgenetically

relevant information.1 The other 90% contain sequences that are highly repetitive and

polymorphic,meaningthattheyarealsohighlyvariable.1Onetypeofthesepolymorphic

sequencesisknownasshorttandemrepeats(STRs),arepeatedsequenceofDNAthatis

usually aroundone to six basepairs in length.2 STRsdiffer in termsof their sequences

betweensitesalthoughwithinasitethenumberofrepeatsoftheuniquesequencediffers

withseriesthatcanbeupto100nucleotidesinlength.2ThesepolymorphicareasandSTRs

are of great interest in the field of forensic science, as they are variable between

individuals.

InForensicScience,STRtyping isusedto infer individualizedsequencingofDNAwhere

todaykitssuchasPowerPlex21andGlobalFilerareusedwithinforensic laboratories in

Australia. Previously kits such as ProfilerPlus have been used, a kit that tests for ten

individual STR loci, although the more STR loci that are tested for, the greater the

discriminationvalue.1ThePowerPlex21kit,asthenamesuggests,testsfor21STRlociand

theGlobalFilerkittestsfor24STRlocibutcurrentlyisonlyusedinonestateofAustralia.

AllSTRlocichosenfortestingwithinkitsaremostlytetranucleotiderepeats(fourbasepairs

inlength)andarecodominant,meaningthattheyareinheritedjustlikeanyothergene,

onefromthepaternalsideandanotherfromthematernalside.1STRsarethereforeable

tobeusedforfamilialsearchingduetothiscodominantinheritancepattern.

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Sinceitsfirstuseinthecourtsystemsin1986,DNAhasbeenahighlyregardedpieceof

evidence, something that has been very hard to argue against. AsDNAbecomesmore

understood,scientistsareabletogainagreater insight intowhohasbeenpresentata

crimescene.IncreasedsensitivityofkitsallowsforensicscientiststogainDNAprofilesfrom

the smallest of biological substances. Increased sensitivity of kits and a greater

understanding overall, has allowed claims such as the secondary transfer of DNA. An

increasedsensitivityofkitsallowsmultipleindividualstobewithinamixtureandagreater

understandingallowsthepossibilityofthesecondarytransferofDNA.Thismeansthatno

longerdoesanindividualdeposittheirDNAontoanobject,itistransferredthroughthat

object(vector)toafinalobject.Thisisbecomingincreasinglycommonwithincourtanda

need for a concise guide to determine the possibility of secondary transfer has now

emerged.AproposedmethodtoproduceaconciseguidetothesecondarytransferofDNA

isthroughameta-analysis,ananalysisthatpoolsresultsfrommultiplestudiestogetherto

increaseitsstrengthandsamplesize.3Theaimofthisliteraturereviewwastoproducea

combined review into the secondary transfer of DNA in addition to drawing multiple

studies together to attempt to provide an analysis of the literature. It is therefore

hypothesizedthatthroughtheexplorationofliteratureandcompletionofameta-analysis

that an appropriate and concise guidewill beproduced todetermine the chancesof a

secondarytransfereventoccurringandwhatconditionsarerequired,toaidBiologistsin

experttestimony.

1.1THETRANSFEROFDNA

TraceDNAhasbeen regardedasa largephenomenonover thepast coupleofdecades

purelyduetoitssignificanceinthefieldofforensicscience.Ithasallowedthedetectionof

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individualsDNAoneverydayobjectssuchasdoorhandles,phones,tablesandchairsjust

basedonasingletouch.TraceDNAisdefinedascellsofftheskinsurfacethathavebeen

carriedontoanothersurfacethroughphysicalcontact.4Thismeantthatfingerprintswere

nolongertheonlywaytoidentifyanindividualatacrimescene,collectingDNAprovided

anewavenueforidentificationbetweenindividuals.TheconceptoftraceDNAwasvery

limitedwhenitwasfirstdiscoveredbutastechnologyhasdeveloped,understandingofit

hasevolved.TraceDNAwas firstexplored ingreatdepthbyvanOorschotandJones in

1997,wheretheyfoundthattheycouldobtainDNAprofilesfromswabbingobjectsthat

wereregularlyhandledbyaparticularindividualandthoseobjectsthatwerepre-cleaned.5

Even though traceDNA isnowrelativelywellunderstood, thereare still somediffering

opinionsastowhereitoriginatesfrom.6-9 ItwasfirstthoughtthattraceDNAoriginated

fromtheoutermost layerof theepidermis,butmore recently studieshave shown that

there isactually littleDNA leftby the time thekeratinocytes (epidermal cell) reach the

epidermis.9Besidesthis, it isnotknownastowheretouchDNAoriginatesfrombutwe

havenowwell-understoodthetechnologiesusedtotestit.AtthebeginningsoftouchDNA

approximately1ngofDNAwasneededtogainafullprofilebutcurrentlyapproximately

only100pg isneeded.10,11Duetothis increase insensitivityof thetechnology,concepts

suchassecondaryandtertiarytransferofDNAhavenowdeveloped.

ThesecondarytransferofDNAinvolvesthetransferofgeneticprofilesfromanindividual

throughavectorandthentoanotherindividualorobject.12Itisdefinedasthetransferof

DNA from one person or object to another via an intermediate person or object, not

involvingadirectlinkbetweenanindividualandthetargetsurface.11,13,14Thefirstinstance

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ofthesecondarytransferofDNAoccurredinastudyconductedbyvanOorschotandJones

in1996intotheprimarytransferofDNA,fromanindividualtoanobject.5Theyswabbed

regularly used objects of individuals and all had provided genetic profiles that were

consistentwiththeowneroftheobject,howeveronetelephonedisplayedageneticprofile

ofaseconduser.5ThiswasthefirstinstancewherethesecondarytransferofDNAwasseen

tooccur.Further,itoccurredthroughanindirectmannerasitwasnotspecificallytested

for,promptingadditionalresearchintothearea.

With continued and more developed research into certain areas also comes with an

increase in knowledge. Even though we are still trying to understand how secondary

transferoccurs,theconceptsoftertiaryandevenquaternarytransferhavearisen.Aslike

secondarytransferofDNA,tertiarytransferinvolvesthetransferofgeneticprofilesfrom

one person or object to another through two intermediary steps involving a vector.15

Quaternary transfer involvesanadditional transferevents, resulting in four transfersof

DNAintotal.Thesephenomenon’shavebeenindicatedtooccurinnumerouspublished

papers,buthavenotyetbeenconclusivelyshowntooccurconsistently.16-20

1.2META-ANALYSIS

InordertocombinestudiesthatexplorethesecondarytransferofDNA,anappropriate

andeffectiveinvestigationmustbecarriedout.Thechosenmethodofcarryingthisoutwas

ameta-analysis.Ameta-analysis is a systematic review following certain criteriawhere

results are pooled and analysed quantitatively.3 They aim to summarise results of

numerousstudies,effectivelyincreasingasamplesizeandthusthepowerofthestudy.3,21

Rareoutcomesofastudymayalsobeovercomethroughtheuseofameta-analysisasthe

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samplesizeofstudiesisincreasedandthereforearareoutcomebecomesanoutlier.21To

carry out a correctmeta-analysis, it should be done as a part of a systematic review,

includingfeaturessuchasacomprehensive,reproduciblesearchandtransparentselection

criteria.21Therearetwomaintypesofreviews,systematicandnarrative.Narrativereviews

exploremorebroader topicsandqualitatively summariseevidence,whereassystematic

reviewsfocusmoredeeplyonprimarystudieswhoexploreclinicalquestions.21

Meta-analyseswerefirstknownbackinthe17thcenturywhenastronomersthoughtthat

thecombinationofdatamightbepreferredtoindividualchoice.22Inthemedicalfield,the

firstcombinationofdata fromnumerousstudiesoccurred in1904,butnotas regularly

usedinthefieldasitwasineducationandpsychology.21,23Thetermmeta-analysiswasfirst

devisedin1976byapsychologistnamedGeneGlass,andnowhasbecomeoneofthehighly

citedpublicationtypeandwhoseuseisstillincreasing.24,25

Recently,meta-analysesaremainlyusedinthemedicalfieldpurelyduethelargeamount

ofinformationthatisgatheredthroughstudies.3Theyaimtoprovidereaderswiththemost

currentresearchpublishedinaparticulararea.3In2007,asearchof“meta-analysis”would

yield1,473articles.3Meta-analyseshavealsobeenusedintheagriculturefieldwiththe

firststudycarriedoutbyFisherintheearly1900’sinvolvingtheprobableandrealconcerns

thatfertilizereffectswillvarybyyearandlocation.26Theyhavealsobeenusedinother

fieldssuchaspublicpolicy,clinicalpractice,psychologyandeducationalresearch.

Twoofthemainpurposesastowhymeta-analysesarecarriedoutaretosummariseresults

of several studies in addition toovercoming small sample sizes in studies.21 By pooling

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theseresultstogether,statisticalpowerofthesestudiescanbeincreasedwhenotherwise

impossible.21Inthemedicalfieldinparticular,performingameta-analysiscanoftenhelp

to reduce “negative” results, helping in providing effective treatmentswithout delay.21

Althoughperformedindirectly,meta-analysismayalsoevenindicateoflackofevidencein

particularareas,highlightingtheneedtoperformmorestudies.21

Although meta-analyses have shown to be a powerful tool, misleading results can be

createdwhenconditionsarenotmetcorrectly.3AsstatedbyWalkeretal.,thereisbelieved

tobefourcritical issuesthatneedtobeaddressedwhencarryingoutameta-analysis.3

These include: heterogeneity of results, availability of information, identification &

selection of studies and the analysis of data.3 For example, during the selection and

identificationofstudies,itmustbeensuredthatbiasinanywaydoesnotcomeintoplay.

Inregardtotheavailabilityofinformation,somestudiestendtoonlyincludesummariesof

theirresults,notrawdata.3Thiscanlimitthetypeofanalysesandconclusionsthatcanbe

madewithameta-analysis.3

Therearetwotypesofmodelsthatareusedtoaccomplishmeta-analyses,randomeffect

orfixedeffectmodels.21Thefixedeffectmodelassumesthatthe‘true’effectsarefixed

whereasa randomeffectmodel assumes thatonlya randomsample is included.21 The

randomeffectmodelismorepreferredforameta-analysisasitbetterreflectsrealityas

studiesbarelyevermimiceachother.21Thisisthereforewhyarandomeffectmodelwill

be used for themeta-analysis carried out in this reviewwhere itwill be attempted to

summariseresultsfromstudiesofthesecondarytransferofDNA.

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2. LITERATUREREVIEW

RegardinganytransfereventofDNA,thewayinwhichitisperformedisrelativelysimilar

acrosstheboard.Theremustalwaysbeaprimarytransfereventinvolvingthemovement

of genetic profiles most commonly from an individual to either an object or another

individual.Thentheremustbeanothertransferevent,hencethetermsecondarytransfer,

wherethatsamegeneticprofilewillbetransferredeithertoanindividual(comingfroman

object)oranobject(comingfromanindividual).Itcannotbesaiddefiantlyastowhichtype

ofmethodwouldbemorecommoninreal-lifesituationsasbothareasfeasibleaseach

other.Intermsofanindividualbeingavector,thismayoccurwhentwoindividualscome

intocontactwitheachother,eitherthroughahandshakeormoreintimatecontact,and

thenthesecondindividualdepositsboththeirownDNAinadditiontothefirstindividuals

onanobjectperhapsatthesceneofacrime.Theothermethodmaymeanthatthetwo

individualsnevercomeintocontactwitheachother.Inthiscaseitmaybemorefeasibleto

callthesecondindividualavictimtoaphysicalcrime.Inthiscasethefirstindividualmay

have touched an object in a public place and the victim had also touched this object

afterwards.DNAtestsonthevictimmayshowtheDNAprofileofthisfirstindividualplacing

thematthesceneofthecrime.

Inthiscase, itcouldbehypothesizedthatthemorecommonscenariothatwouldoccur

would be that of an individual as the vector. Often in the course of an investigation,

individualswillbelinkedwiththecrimebasedontheirDNAbeingatthescene.Inthiscase,

itcouldbearguedthatsecondarytransferwasthecausefortheirDNAbeingpresentatthe

crime scene especially if they had never visited that area. Below studies using both

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instancesasvectorswillbeexploredandreviewedtogainagreaterunderstandingasto

howthismechanismoccursanditslikelihood.

2.1 OBJECTASVECTOR

When the secondary transfer of DNA is put forth as a possible explanation, it ismore

sensibletohaveoccurredthroughanobjectratherthananindividual.Thisscenarioatleast

isbelievedtobemorereasonableinhowitoccurs,asthechancesofthetwoindividuals

coming intodirectcontactwitheachother ishighlyunlikely.Oneofthefirststudythat

performedsecondaryDNAtransferthroughanobjectwasperformedin2012.Theauthor’s

usedacasescenariotobasetheirexperimentsoff.Thefirstexperimentfollowedhands

comingincontactwithakid’stoyorasingletandthenthatobjectcomingincontactwith

a laboratory coat, both actions occurring for one minute and immediately after each

other.27 The results showed that in 19 out of 20 repeats, DNA from the donor was

observed.27Thisstudyhoweverdoesn’tmimicreal-lifenearlyasmuchasthecasestudy

wouldhave.Actionswereperformedimmediatelyaftereachotherwithnotimedelayand

theindividualwhoparticipatedinthesecondtransfereventwaswearinggloves,therefore

notcreatingtheissueofadditionalDNApresent inthetransferevent.This increasethe

transferpercentagelikelihoodasthedonor’sDNAistheonlycellularmaterialpresent.

ThefindingsinGorayetal.arealsoinaccordancewithastudyconductedbyLehmannet

al.thefollowingyearwheretheyadditionallyobservedsecondarytransferoccurring.27,28

Lehmannetal.additionallyusesglassasavectorobject,howeverunderwentatransfer

eventontothesametypeofmaterialasthevector.28Similartothestudyconductedby

Gorayetal,thisstudydoesnottakeintoconsiderationofbackgroundDNAthatmaybe

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presentthroughtransfereventstherefore increasingthepercentageofcellularmaterial

transferred.Incontrast,Verdonetal.conductedanexperimentthesameyearthatused

sevendifferenttypesofsubstratesasthevectororsecondarysubstrateandusedeither

plastic or cotton as the opposite substrate.29 The differing types of substrates paired

againstoneanotheryieldedthesameresultofthatinLehmannetal.,wheresecondary

transfer had shown to occur. However, within the study there were differences seen

betweentheporosityofthesubstrates.Whenthetransferwentfromanon-poroustoa

poroussubstrate,thetransferofDNAwashigher.29

Auniqueapproachwasusedby Frenchet al.whereUltraviolet (UV)powderwasused

throughthetransferexperiment.Thisapproachcanbeusedformultipletypesofmaterial

thatmaybetransferredthroughanindividual’shand.Threeparticipantssataroundatable

drinking from a bottle with their own three glasses for 30 minutes where only one

individualstartedwiththeUVpowder.30Therewasnodirectcontactbetweenindividuals

butthestudyshowedtheindirecttransferoftheUVpowdertoallindividuals.30Obviously,

UVpowderdoesnotmimictheactionsofDNApreciselybutitmaybeusedasabeginning

point in order to understand the mechanisms behind the transfer especially within

networksofindividuals.

GorayandvanOorschotperformedaverysimilarexperimenttoFrenchetal.thefollowing

yearwherethesame“uncontrolled”conditionswereused.Herehowevertheydidnotuse

UVpowderbuttestedfortheDNAitself.Intheareaimmediatelysurroundingparticipants

from the chairs and table, no other DNA than that of the relevant participant was

detected.31 In27%of the table samplesand then33%of thechair sampleshadaDNA

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profilesfromparticipantsthatdidnotcomeintocontactwiththesurfaceindicatingthat

theseprofilesmighthavebeentransferredfromthejug.31Afurther58%oftablesamples

and42%ofchairsampleshadDNAprofilesfromunknownindividuals,possiblyduetothe

secondary transfer of DNA.31 These results support the findings of French et al.where

secondarytransferdoesoccurthroughaseeminglynormalsocialsituation.Onedownside

tothestudyconductedbyGorayandvanOorschotwasthattheyhadcleanedmostareas

involved in the experiment beforehand meaning that background DNA would not be

present. In this case transferpercentagesmaybeuntruly inflatedwhich is the sameof

previousstudies.28-30

In2015, researches seemed to realise the idea that secondaryDNA transfermayoccur

withinthelaboratorysothereseemstobeaninfluxofthestudiesconductedwithobjects

frequentlyused in laboratoriesactingas thevector.Margiottaetal.usedglovesas the

vectorfortransferwhere16usedgloveswereexaminedforDNApresent.Inhalfthecases

tested for, they found amixture of DNA relating to both the test samples and alleles

belonging to an unknown individual different to that of the volunteer.32 In addition, in

another15%ofthecasesjustallelesfromanotherunknownindividualwerefound,unable

tobeattributedtothevolunteer.32Thiscouldbeattributedtosecondarytransferbythe

volunteerontotheirglovesfromanunknownsourcealthoughauthorsdidpointoutsome

oftheirnegativecontrolsfromsupposedly“clean”glovesdidyieldallelespresent.32

Szkutaetal.continuedonfromotherstudiesthatfocusedontheamountofDNAthatis

leftonvectorsafter the transfereventhasoccurred.Theydid thisby theconductinga

secondary transfer event of DNA between two “exhibits” using tools such as forceps,

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scissorsandglovesasthevector.33Itwasshownthatsecondarytransfercouldoccurfrom

exhibit to exhibit byDNA-free vectorsbut themain findingwas still having a sufficient

quantityofDNApresentonthevectortocontinuetoproducesecondarytransfertoother

objects.33ThesefindingsareparticularlymoreprevalentforbloodratherthantouchDNA

during the transfer event which seems to be due to the higher concentration of DNA

present,additionallyshowninnumerousotherstudies.28,34-36Herethoughthisstudyused

DNA-freevectorswhichagaininreal-lifescenariosisveryunlikelytooccur.Asmentioned

previously,thismayincreasethetransferpercentagehencegivingfalseindicationsofthe

secondarytransferofDNA.

Fonneløpetal.investigatedthetransferofDNAfromanobjecttoindividualandthenback

to an object, with this idea coming fromMeredith Kerchermurder casewhichwill be

furtherexploredbelow.Theyusedthreevolunteerswhowereknowntobegoodshedders,

thosethatdepositafullDNAprofile15minutesafterhand-washing.17Thefirstindividual

pickedupanobjectandhandleditfor30secondsthenplaceditback.17Asecondindividual

thenhandledthissameobjectandthensubsequentlyputpressureonapieceofbench

paperorfabricwithpressurecausingatertiarytransferevent.17Substratesincludedinthis

studywereplasticconicaltubes,metaldoorhandle,disposablegloves,fabricandbench

papers.17Results showed for the first transferevent,83%of theDNAwashighenough

quality for thedetectionof a full profile and for the second transfer event, that figure

droppedto53%whichsawthoseprofilesuptothestandardofcasereportinganddatabase

searches.17

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In2017,Buckinghametal. followedon from their study in thepreviousyearandused

cottoncoveredglassplates(porous)ratherthanjustglassplatesthemselves(non-porous)

toinvestigatetheresultingyieldandprofileofDNA.Theyagainusedthesamemethodof

theknifeasthetransfervectorwiththefourindividualsalltouchingitatonestagethen

placingtheirhandonthecottonplate.3740handprintswerecollectedwithsomeresults

producingevidenceofsecondaryandtertiarytransferofDNAalthoughtheproportionof

thistothedepositor’sDNAwaslessthan10%.37Therewereadditionallysomeunknown

sources of DNA found in swabs from cotton plates.37 In comparison between both of

Buckingham et al.’s studies show that the transfer of DNA both directly and indirectly

appearstobedependentonthesurfaceinwhichtheDNAisrecoveredfromaswellashow

theDNAisrecovered.37

2.1.1LAUNDERING

A laundrymachine for thetransferofDNA isa relativelywellcoveredtopic in termsof

spermatozoabutthisstudyconductedbyKamphausenetal.in2015wasoneofthefirst

testingfortouchDNA.15individualsrubbednecksagainstacloth(primarytransfer)for5

secondswithmediumpressure.35Witha totalof46 independentwashescompletedby

handormachinewith/withoutdetergentsecondarytransferoftouchDNAwasdeemed

almostimpossibleforreliableSTRanalysis.35

In 2018, another studywas conducted into themechanismsof the transferofDNAvia

laundry.Eightunwornsockswerewashedinanormalloadofhouseholdlaundryoffour

families(2perwash)andthensubsequentlyair-dried.20Resultsshowedthatoutofthe32

samplesthatwerecollectedafterlaunderingonly7ofthem(22%)yieldedaresulthigher

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thantheminimalthresholdforcasework(0.06ng/µL).20Fourofthosesamplematcheda

profileofafemalefromthehouseholdwhichwaseitherasinglesourceormajorprofile

indicative of the transfer ofDNAoccurring but unknown as to either direct or indirect

transfer.20

2.1.2FINGERPRINTBRUSHES

MoststudiesconductedexploringthesecondarytransferofDNAinvolveavectorthatis

mostlikelyanobjectsuchasaglassbeakerortube,butthereareobjectsthatareusedin

theprocessingofcrimescenesthatmaycauseforexploration.Fingerprintbrushesareused

byForensicScientistsandPoliceOfficerstodeveloplatentfingerprintsatthesceneofa

crime.Oftentheseindividualsdonotusedisposablebrushesbutmorecommonlyreusable

brusheswithoutthinkingofbiologicalcontaminationissuesthatmayarise.VanOorschot

etal.wasthefirsttotesttheextenttowhichDNAtransfermayoccurusingsquirrel-hair

fingerprintbrushes.38Outof26samplesfrompre-usedbrushesbrushedonpaper,only2

resultedinprofileswhichwerepartial,bothbrusheswereusedincasework.38Another73

squirrel-hair fingerprint brusheswere in current use by staff andwere brushed over 5

sheetsofplasticwith4beingcleanand1(the3rdinsequence)presentingafreshdeposited

handprint.38TheresultsshowedthatverylimitedpickupandtransferofDNAoccurredby

thebrusheshoweverwhenchangesweremadetotheDNAanalysis,significantpickupand

transferofDNAwasseen.38

ThepreviousstudyhadconfirmedthatthetransferofDNAcanoccurthroughfingerprint

brushesatcrimescenesbutnotsufficientlytoshowthatitoccursconsistently.Asecond

studywasconductedbyProffetal.wheretheydeterminedthatDNAcontaminationwith

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fingerprintbrusheswasquitecommon,where85%ofthebrushestheytestedresultedin

eitherfullorpartialprofilespresent.39Intestingbrushesforthepotentialforsecondary

transfer,theauthorsused16brushesagainstanacetatesheetwithcarbonblackpowder

with25%ofsurfaceshadDNAdetectedwithonefullprofile.39Aswiththepreviousstudy,

this onealso founda limited riskof the secondary transferofDNA through fingerprint

brushes.38,39

Even though these are the only two studies that have been explored in the secondary

transfer of touch DNA, there have been studies performed with other biological

substances.Thesewillbereviewinlatersections.Fromjustthesetwostudies,itshowsthe

needtobemoreawareofcontaminationandtransfereventsoccurring.Onesetofauthors

event suggest thatdecontaminationprocedures shouldbeput intoplace regarding the

continuous use of fingerprint brushes between crime scenes.39 In today’s crime scene

examination, this has already been put into place where either disposable fingerprint

brushesorde-contaminatedfingerprintbrushesareused.

2.2INDIVIDUALASVECTOR

Although this scenario seems like themore common scenario to occur in a “real-life”

situation,itseemstobetheleastresearchedofthemall.Thefirststudythatdidusean

individualasthevectorforthetransferofDNAwasconductedbyLoweetal.in2003.The

authorsusedpairingsofgoodandpoorshedderswhoshookhandsforone-minuteand

then the poor shedder gripped an object for 10 seconds.40 One pairing in particular

consistentlyfoundthatthegoodsheddermorethanoftenhadtheirfullprofiletransferred

andthepoorshedder,thevector,couldnotbedetected.40Byputtinginplacea30second

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timedelaybetweentransferevents,mixedprofileswerediscoveredwhere70%werefrom

theoriginalsource.40AlthoughthisstudydidconfirmeventsofsecondaryDNAtransfer

occurring, it is performed under controlled conditions where pairings were created to

optimizethedesiredoutcomeandtypicallytheremaybehoursordaysbetweentransfer

eventsoccurring.

Farmenetal.additionallyconductedastudywheretwoindividualsundertookahandshake

for30secondsfollowedbythegrippingofbeakersfor30seconds.14Atotalof60swabs

weretakenandallresultedinaDNAprofileofsomeform,eitherpartial,fullormixed.14

Althoughnotspecified,theauthorssuggestthatallsamplesweremixturescontainingboth

profilesoftheknowncontributors,indicatingthatsecondaryDNAtransferhadsuccessfully

occurred.14ThisisinagreeancewiththepreviousstudyconductedbyLoweetal.where

theyalsosawsecondarytransferoccur,withbothstudiesseeingthatmixturescanhave

highercontributionofindividualswhowerenotthevector.14,40Typically,inthescenarioof

secondaryDNAtransfer,theindividualwhodepositstheDNAatthelasttouchwillleavea

higheramountoftheirDNAasthattransferisprimary.ThoseDNAprofilesthathavegone

throughmultiple transfer before the final depositwill typically decrease significantly in

concentration.

AstudyconductedbyMeakinetalin2015alsoshowedinstancesofthesecondarytransfer

ofDNAoccurringthroughahandshakeasthevectorandaknifeasthefinaltransferevent.

A“regularly-used”wasartificiallycreatedoveraperiodoftwodaysandafterthattime

period two volunteers shook hands then immediately stabbed their knife into foam

occurringforthenextthreedayswiththreedifferentknives.41Thestudyfoundthatfor

20

threeofthefoursetsofknivesmixtureswerefoundcontainingthreedifferentprofiles.41

Thefirsttwocouldbeattributedtotheregularuseroftheknifeandthehandshaker,but

the other was an unknown profile, possibly the cause of secondary transfer from the

regularuserthroughothermeansorpossiblytertiarytransferfromthehandshaker.41As

with the study conducted by Lowe et al., this study has tested the persistence of the

secondarily-transferredDNAandfoundthatthebestchanceofrecoveringthatDNAwould

be to sample as soon as possible after the final transfer event had occurred which is

consistentwithLoweetal.’sfindings.40,41

Withanew-foundgap inthe literature,Caleetal.performedastandardstudy intothe

secondarytransferofDNAwithakitofincreasedsensitivity.Theauthorsaimedtostudy

howthepresenceofsecondaryDNAtransferwouldaffecttheinterpretationofDNAtyping

results.11Inaddition,theeffectsbetweensurfacetexturewasalsoexplored.11Participants

washed hands before wearing glove for 1.5 hours before shaking hands with another

participantfor2minutesandthenhandlingapre-cleanedknifeforafurther2minutes.11

Significantresultsofthisstudyconcludedthattexturedidnotappeartohaveasignificant

effect in addition to the data obtained from five of the 24 samples that suggest that

individualscanhavetheirDNAdepositedonobjectsinsufficientquantitiesthattheycan

be identifiedas theonlyormajor contributor, suggesting that theyhad come indirect

contactwith the object.11 These results seem to be a trend through numerous studies

whereifgoodsheddersarethedonorDNAprofile,theywilltypicallyoverpowertheDNA

of that of the vector.9,42-44 All these papers discussed within this section are highly

unrealisticandlackscientificrigour.Inaneverydayscenario,individualswouldbarelyever

21

shakehands for30 secondsand furtherhandleanotherobject immediatelyafterwhen

committingacrime.

2.3INDIRECTFINDINGSOFSECONDARYTRANSFER

TherearealsoinstancesinwhereprimarytransferorpersistenceofDNAmayhavebeen

experimentedonduringastudyalthoughunknownallelesmaybedetectedontheobject

or individual.Hereall the studies thathavedetectedsecondary transferofDNAduring

anotherexperimenthavebeencollated.However,inthesestudies,unlesstheauthorshave

specificallystatedwedonotknowwherethesecondarytransferhasoccurredandwhothe

unknownallelesarefrom.Additionally,theunknownallelesmaybepartofthebackground

DNApresentonanobjectunlesstheobjectofindividualhasbeenpre-cleanedbeforehand.

ThefirstinstanceofaccidentaloccurrenceofsecondaryDNAtransferwasduringastudy

conductedbyLoweetal.in2002wheretheauthorstestedshedderstatusesofindividuals.

Secondary transferwasdetectedwhenparticipantshelda tube for10 secondsbut the

extent was dependent on the subject pairings.42 In an additional two experiments,

secondary transfer was not conclusively shown as DNA recovered varied considerably

betweenreplicates.42Overall,theauthorsconcludethatthesecondarytransferofDNAwas

showntooccurunderidealconditions.42Afewyearslater,Phippsetal.conductedastudy

ingreatsimilaritywheretheyfoundthattheoccurrenceofsecondarytransferwaslowand

notlikelybutpossible.43

AstudyconductedbyRuttyfoundaninstanceofpossiblesecondaryDNAtransferoccurring

wheremale-female pairs were used in order to simulate a strangulation event.45Men

22

placedtwofingerpadsonthewomen’sneck,anactionthatwasrepeatedatotalof29

timescreating116samples.45Twoswabsofthemale’sfingers(controls)yieldedaprofile

ofafemaleinthestudyalthoughthesetwoindividualshadneverbeenpairedorcomeinto

contactwithoneanother.45Partialprofileswerealsofoundfromoneormoreindividuals

in pairs of controls up to 10 days after the experiment.45 Although in a different

circumstancetothatofLoweetal.,bothstudiesshowthepresenceofadditionalalleles

thatmaybeattributedtosecondarytransfer.

Usinganotherdifferentexample,Petricevicetal.usedfivevolunteerswhosleptintheir

ownbedbutwithanewlowerbedsheet.46Anotherscenariosawthatindividualssleptin

a bedwhere they never had previously.46When individuals slept in their own beds, a

second individual’s DNA profile was found in at least one sample from 3 out of the 5

volunteers.46 The authors were able to attribute these from the volunteer’s partners,

possiblytransferrinfromotherbeddingusedorfromthevolunteerthemselves.46When

individuals slept in an unknown bed, all unknown samples could be attributed to the

regularuser’sprofilewhichdidnotindicatesecondaryDNAtransfer.

AnothersimilarexperimenttothatconductedbyLoweetal.andPhippsetal.,Djuricetal.

hadvolunteersholdplastictubesandindividual’sanklesfor10seconds.47Whenvolunteers

heldtubes,outofthesevenprofilesobtains,oneresultedinapartialprofilenotmatching

thevolunteer’s,indicatingtheoccurrenceofsecondarytransfer.47Whenankleswereused

asdepositionsites,nooccurrenceofthesecondarytransferofDNAwasnoted.47Gorayet

al.conductedaverysimilarexperimentagainbuttestedthedepositionontoglassplates

ratherthanplastictubes.Atotalof40samplesweretakenandnon-selfDNA(unexpected

23

alleles) was seen in 79% of the samples.9 Surprisingly, 7 samples had the primary

contributorprofileexcludedfromthemixture.9Gorayetal.hypothesizedthatthemost

commonexplanationforthepresenceofnon-selfDNAindepositswereduetothetransfer

ofnon-selfDNApickedupthroughevery-dayactivities.9Theauthorsalsoranthemixtures

through staff eliminationdatabasesandotherworkerswhowereknown tobe in close

proximitytoparticipantsmatchedtothenon-selfDNAdeposited.9

ApaperpublishedbyOldonietal.appliedtheconstantunknownfromcontactstainsat

break-instoconductedanexperiment.Here,volunteerswouldhandleanobjectregularly

over 8-10 days then a second userwould handle the same object for a pre-determine

period of time.48 A total of 231 samples were created where approximately 2-10%

containedunknownalleles,possiblesuggestingsecondarytransfer.48Additionally,intwo

ofthesimulations,therewerefullDNAprofilespresentthatdidnotcorrelatetoindividuals

directlyinthosesimulationsbutratherothervolunteersfromthestudy.48

Manyof thesestudiesconducted in thesecondary transferofDNAarecarriedoutata

forensic laboratory and involve volunteers who work in an office/laboratory based

environment. As these individualsmay be at high risk for transferring DNA to forensic

exhibits if working with them, an exploration into the ways in which volunteers may

transfer DNA between themselves was conducted by Fonneløp in 2015. Here four

volunteershadtheirkeyboardsandmouseswappedwithanother’swithswabstakenfrom

bothobjects andvolunteers at thebeginningand throughout the study.49 Swabs taken

before the study had begun showed DNA present from unknown individuals, but

unfortunatelyitcannotbeconcludedifthisDNAwassecondarilytransferredorthrough

24

directcontact.Theauthor’sattributedthisoccurrencetobackgroundDNAbeingpresent

oneverydayobjects.49Inaddition,thestudyalsoshowedthatsecondarytransferofDNA

occurredthroughthepresenceoftheoriginaluser’sDNAonthehandsoftheseconduser

upto8daysafterreceivingthecomputerequipment.49

Oneof the simplest ofways todetermine thepossibility of secondaryDNA transfer or

backgroundDNApresentwouldbetodirectlyswabanindividual’shandwhichwascarried

out by Lacerenza et al. 120 sampleswere directly collected from60 individual’s hands

where56sampleshadoneormoreunknownallelesand36ofthosecouldbeclassifiedas

mixtures(64.3%).50AstheseDNAsamplespresentinthemixtureswereforeigntothatof

thevolunteer,itcouldbeconcludedthateithersecondaryDNAtransferoccurredorthat

theseprofileswereapartofthebackgroundDNApresentonthatindividual’shand.50

AnewsubstratewasthenusedbyvandenBergeetal.where individualsdraggedeach

othermimickinganactivity-relatedscenario.Samplesweretakenfromtheknee-areaof

thetrouserswherethedraggerdidnottouchresultingin26samples,3ofwhichdidnot

resultinover7alleles.51Donoralleleswerepresentin100%ofthesamplesandnon-donor

in71%.51Theauthorsconcludedthatmorecaremustbetakenwheninterpretingmixtures

asallelesmatchingaperpetratormaynotbedistinguishablefrombackgroundsignals.51

Inaveryrecentstudy,Mageeetal.usedcollarsandcuffsofuppergarmentstodetermine

the presence of DNA.52 Out of 55 samples taken, non-wearer DNA was recovered, an

averageof1.3ngfromthecollarand2.7ngfromthecuffs.52Fromoneparticularsamples,

anon-wear contributedapproximately57%ofDNA toamixture, followedby thewear

25

contributing37%andasecondnon-wear,7%.52Threeothernon-wearersampleswereable

tobeattributedtospousesofthevolunteers.52

2.4SHEDDERSTATUS

Shedderstatusisoneofthemorerecentconceptsundertheumbrellaofthetransferof

DNAandonethatwillcontinuetogrowasmoreresearchisconductedintothetopic.The

first study to directly carry out an experiment on individual’s shedder status’ was

conductedbyLoweetal.in2002andisconsideredthebackbonestudytothistopic.The

authorsgotparticipantstogripasteriletube,15minutes,2hoursand6hourperiodsafter

hand-washing.42 They determined that there were differences between individuals in

regardtohowmuchDNAthattheydepositedonasurface.42Goodshedderswerethen

classediftheindividualleftanentireprofileafter15minutesofhand-washing.42Allother

individualswereclassedaspoorshedders.

Afair fewyears later,Phippsetal.conductedasecondstudythat investigatedshedder

statusandusedalmostidenticalexperimentstothatofLoweetal.Plastictubeswereused

for deposits and hand washing occurred 15 minutes prior to the experiment.43

Unexpectedly,thestudyhadshownthatindividualsdonotproduceconsistentquantities

ofDNAovertime,varyingasmuchwithinthemselvesthancomparedtootherpeople.43

Finally,usingthecriterialaidoutinLoweetal.,thisstudydiscoveredno“good”shedders

andallwereclassifiedas“poor”shedderssuggestingthislevelofclassificationmaynotbe

allassimpleasoncethought.42,43

26

ThefollowingyearFarmenetal.conductedastudyonshedderstatusinconjunctionwith

anexperimentonsecondaryDNAtransfer.Authorsmeasuredindividuals’shedderstatus

justbasedonswabsfromtheirhands.44Thisunfortunatelydoesn’ttestthequantityofDNA

thattheindividual’sactuallydepositonasurface,notclassingasatruerepresentationof

shedderstatus.Outofthenineindividualswhoparticipatedinthestudy,twoindividuals

had a low DNA yield and seven individuals produced profiles that were able to be

analysed.44

Morerecentlyin2016,Gorayetal.carriedoutanexperimentthatexploredtheeffectof

thevariableshanddominance,handsizeandgenderonindividual’sshedderstatuses.240

sampleswerecollectedfromhandprintsonglassplateswhereonlyfourofthesampleshad

nodetectablequantitiesofDNA.9 Twoofthetenparticipantscouldbeclassedasgood

sheddersbutthiswasdoneunderdifferentcriteriaandexperimentalcriteriaastothebase

studyconductedbyLoweetal.9Theonlycorrelationorsignificancethatthestudyfound

with the variables tested was that of gender, where a significant difference was seen

betweengendersintheamountofDNAthattheydeposited.9

2.5OTHERFORMSOFDNAINVOLVEDINSECONDARYTRANSFER

2.5.1TRANSFEROFSPERMATOZA

Oneof the first studiescarriedout in thebroader realmof the transferofDNAwasby

Kafarowskietal.in1996.Thiswasconductedbeforethefirstoccurrenceofthesecondary

transfer of touch DNA andwas one of the first of its kind. It aimed to determine the

likelihoodofthetransferofspermatozoabetweenarticlesofclothingduringamachine-

cyclewash.53Althoughtheretentionofspermatozoaduringmachinewashhadpreviously

27

beenexplored,thetransferofittootherarticlesofclothinghadn’t.54,55Theauthorsused

underpants as a means for the primary deposit of spermatozoa, reflecting real-life

situationsthatmaydepictasexualassault.Thispairofunderpantswaswashedwiththree

additional clean pairs and then machine dried.53 In all three trials, trace quantities of

spermatozoawerefoundwhereaminimumofonespermheadandmaximumofeightper

sample.53AsthiswasoneofthefirststudiesperformedinthesecondarytransferofDNA,

there are many downfalls to it that have improved over time with knowledge and

technology.

2.5.2TRANSFEROFSALIVA

SalivaisanadditionalbiologicalsubstancethatcontainshumanDNAthathasthepossibility

tobetransferredthroughobjectsandindividualstocauseasecondarytransfer.Thefirst

thatexploredintothissecondarytransferofsalivawasconductedbyvanOorschotetal.

wheretheyexploredthetransferofsalivathroughasquirrel-hairfingerprintbrush.Inone

experimentasinglebrushwassweptover19sheetsofpaperwithdried-salivapresentthen

brushedoveranother20clean sheetsofpaper.38Outof the80 samplesgenerated,13

resultedinpartialprofiles,1innoprofileandtheremaining66resultedinfullprofiles.38

The second experiment saw both the powder type and DNA extraction tested in the

transferofsalivafrombrushtosixsheetsofplastic.38Forthecomparisononextraction

techniques,outof6samples,3werefullprofiles,2partialprofilesand1noresult.38Any

presenceofaprofileinthisinstanceconfirmsthesecondarytransferofDNAhasoccurred

fromthepaper,tofingerprintbrushandthenbacktothepaper.

28

Nextresearchesdecidedtocarryoutexperimentverysimilarinnaturetothoseconducted

withtouchDNAbutwithsalivainstead.Wiegandetal.transferredsalivaontoeitherpaper,

cottonorplasticsurfaces,leftthemtoairdrythenrubbedathumbontothestrain.34The

thumbwasthenplacedontoanotherpieceofpaperandthentheareawasswabbed.34

Whenpaperwastheprimarysurface,onlyabout50%ofthetransferredsalivaproduced

DNAprofilesalthoughtheywereinverylowconcentrations(max.1pg/µL).34Intotalonly

3outof96gavecompleteprofiles,allfromplasticastheoriginalsourcebutonlyonebeing

fromsaliva.34

Thesecondarytransferofsalivawasadditionallytestedthroughavectorofpensandplastic

tubes for theoverall aimof determining if salivawouldbe amoreprevalent biological

materialduringtransferevents.16Theirstudyconcludedthatsecondarytransferofsaliva

did occur, in greater levels of retention than touch DNA and that moist surfacesmay

facilitateDNAtransfertoagreaterdegreethanadrysurfaceaspreviouslydiscoveredin

Gorayetal.2010.16,56TheauthorsadditionallystatedafindingofbackgroundDNApresent

onanindividual’shandwhendealingwiththetransferofDNA.16

In2015,thetransferofsalivathroughlaunderingwasstudied,somethinginwhichtouch

DNAhadbeenexploredmultipletimes.Individualsdepositedsalivaontotextileclothsand

thentheseweresubsequentlywashedwithcleantextilecloth.35Betweentheexperiment

additionallyconductedwithbloodandtouchDNA,atotalof46independentwasheswere

completed both by hand and machine with/without detergent.35 Saliva was seen to

undergotransfereventswhichsupportsotherstudiesperformed.16,34,35,38,56

29

The most recent study conducted in relation to the transfer of saliva was through

individual’s hands as vectors for the transfer. 14 areas of the individuals palm were

swabbed inaddition to theglassplatewhich the salivawas transferred to.57 The study

showeda significantdecrease in thequantityofDNA through the final transferevents,

wherepreviouslyanaverageof9.16ngwasseenandposttransfertherewasanaverageof

0.225ng.57Thestudyalsoyieldedanumberofunknownallelesonthefinalplatewherethe

authorscontributedthistopriorinteractionwithindividuals,supportingthefindingthat

secondarytransferofDNAhadoccurred.57

2.5.3TRANSFEROFBLOOD

Followingonfromthesecondarytransferofsaliva,Wiegandetal.additionallyconducted

astudyintotheriskofthetransferofbloodstainsfromsurfacessuchasplastic,cottonand

paper.Thebloodstainswerelefttoairdrythenanindividualplacedtheirthumbonthe

stain thenontoanotherpieceofpaper.34Fromthecottonsurface,DNAconcentrations

aftersecondarytransferoccurredwereremarkablysimilartothosefollowingtheprimary

transferevent.34Whengloveswereused insteadofbarehands, theDNAconcentration

followingDNAtransferincreased.34Boththeseresultssupportedthefindingsofsecondary

transfer of DNA through blood occurring. Lehmann et al. conducted a very similar

experimentbutall transfereventswereconductedwith substratesonlyandnohuman

interaction. The experiment supported the existence of the secondary transfer of DNA

throughbloodandadditionallyfoundthatifthebloodwerewet,itwouldtransferfurther

andwouldendinahigherconcentrationasmoresubstancewastransferred.28

30

AstudyinvestigatedsecondarytransferofDNAthroughlaunderinginawashingmachine

orbyhandbyKamphausenetal.in2015.Artificialbloodstainswerealsocreatedontextile

cloth, then dried and these were then washed with clean textile cloths.35 In total 46

independent washes were completed both by hand and machine in addition to

with/withoutdetergent.35 Transferofblood cellswere seen in this study in addition to

numerouspreviousones.28,34,35,36

Agapwasthenidentifiedintheliterature,wherenoneofthesepreviousstudieshadreally

thoroughlytestedtheeffectthearidityofthebiologicalsubstances,herespecificallyblood.

vanOorschotetal.researchedintothisusingcottonasthesecondarysubstrate.36Drying

time, temperature and humidity all acted as dependent variables in this study.36 The

secondarytransferofbloodwasseentohaveoccurredwithsignificantdifferencesinthe

transferpercentageofbloodbetween5and60minutes.36

2.6TERTIARYTRANSFEROFDNAANDBEYOND

Although the secondary transfer of DNA is still in the process of gaining a greater

understandingofitsmechanisms,throughthesestudiesthediscoveryoftransferevents

occurring after a second transfer have occurred. These studies are still in their very

preliminarystagesandlikesecondaryDNAtransfer,stillrequireagreaterunderstanding

intothemechanismssurroundingtheprocess.Thefirststudythatexploredtheoccurrence

ofthetertiarytransferofDNAwasconductedbyWarshaueretal.whereanobjectwasthe

original vector andwas further transferred from an individual to another individual or

objectasthetertiarytransferevent.16TheywereabletoprovethattertiarytransferofDNA

couldoccurbutwereunabletorecoverenoughDNAtoestimatetherateofDNAlostduring

31

the additional transfer step.16 Out of the additional transfer steps conducted, 87.5%

displayed less than half of the expected alleles.16 These results correlated with those

producedbyFonneløpetal.whereafterthefinaltransfersteponly17%ofsampleswere

highqualitythatcouldbereportedandgothroughdatabasesearching(5outof30transfer

events).17

In the study that was conducted by Fonneløp et al. they also tested the instance of

quaternarytransferoccurring,whichwasafirst.Theyfoundthatonceagain,itoccurred

butaswithmostof the issueswiththestudiesexplored inthis review,all surfacesand

objectswerecleanedthoroughlytobeginwithtoeliminatethebackgroundDNApresent.

Atotalof17outofthe108analysedsamplescontainunknownallelesandtheseunknown

allelescouldnotbeattributedtothepresenceofbackgroundDNAasitwaseradicated.17

Therewasadditionallyonetransfersequencethattheauthorswereabletodeterminethe

donoroftheDNApresentinthetertiarytransferevent.17ThesourceoftheDNAwasfrom

thegirlfriendofthevolunteer,asamplefoundinalltransferevents.17Previously,thesetwo

individuals had not been in contact with each other for more than 10 hours and the

volunteerhadadditionallywashedhishandspriortotheexperimentbeginning.17

In2016,Helmusetal.performedthefirststudysinglytestingthetertiarytransferofDNA

duetothegapidentifiedwithintheliterature.Threepairsofindividualswereusedwitha

cotton cloth as the vector and final depository where a total of 180 samples was

generated.19 Out of those 180 samples generated, only 72 had detectable DNA that

identifiedthattertiarytransferhadoccurred(40%).19Inastudycompletedtheyearearlier

theyhadadditionallyshowntheoccurrenceofthetertiarytransferofDNAbuttheauthors

32

notedthattheywereunabletodistinguishbetweentertiarytransferorinterferencefrom

alow-levelofnon-donorDNA,somethingthatmayhavetodowithtypeofsystemused,

thePromegaESI-17FastPCR.18

Inamorerecentstudy,tertiarytransferofDNAevolvedtoamorecomplexmechanism

thanthatofindividual-objectinvolvedtransfer.Sixnewunwornsocksandat-shirtwere

launderedtogetherwithnoadditionalitemsinamachinethathadbeenpreviouslyused.20

ResultsshowedthattherewerenoDNAresultsrecoveredfromquantity(abovethreshold)

throughtoprofilingthereforenotsuggestingthat tertiary transfercouldoccur.20 In this

case,either thekitswith improvedsensitivityneedtobecreatedor this is thepointat

whichtheextentoftertiarytransferwilloccur.Thisishoweverunknownwithoutadditional

studiesconductedinthisarea.

2.7CASESTUDIES

In2002,apaperwaspublishedbyAnsellexploringacasestudyinvolvingthesecondary

transfer of seminal constituents. A rape occurred in Sweden whereby a woman was

allegedlyrapedbyamanwheremixedDNAprofileswerefoundinboththevaginalswabs,

underwearswabsandpenilesamples.58Bothvaginalandunderwearswabsexcludedthe

manas a suspect ashisDNAwasnot found (hada vasectomy), although thewoman’s

boyfriendDNAwasadditionallyfoundduetoconsensualintercoursethepreviousday.58

Thepenileswabsalsoshowedamixedprofilebutthatofthewomen’sDNAinadditionto

thewomen’sboyfriendsDNA.58Theallegedrapeofthiswomenwasabletobeprovedin

courtbythesecondarytransferoftheboyfriend’sDNAtothesuspect.58

33

Morerecently,in2017,anarticlewascreatedfollowingtheincidenceofcontaminationby

PoliceOfficer’sinAustriawhichwithouteliminationdatabases,manyinstancesofindirect

orsecondarytransferofDNAmaygoundetected.59Duringtheperiodof2000to2016,a

totalof46,000tracesamplesweresubmittedthroughDNAandoutofthoseatotalof347

contamination incidents were detected through the use of the elimination database

(0.75%)withamajorityfromthecauseofindirecttransfer.59Theauthorsfurtherwentinto

detailregarding3separateincidentsthattherewereabletopinpointwherethetransfer

eventshadoccurred.Thefirstoccurrencehappenedthroughsharedcameraequipment

that hadbeenusedby a PoliceOfficer completely unrelated to the case.59 The second

showstheindirecttransferbyaPoliceOfficerthroughasharedvehicletoavolumecrime

caseandthethirdthroughpackagingofevidencematerialonanunrelatedPoliceOfficer’s

desk.59

3. PERFORMANCEOFMETA-ANALYSIS

3.1METHOD

Athoroughreviewoftheliteraturewasconductedthroughtheuseofonlinedatabasesto

gatherarticles inthesecondarytransferofDNA.Databaseswerechosenbasedontheir

relevancetothetopicofForensicScience.DatabaseschosenwereScopus,WebofScience

CoreCollection,ProQuest,Medline,ScienceDirectandResearchGate.Somedatabases

weremorespecifictoForensicSciencebutgeneralsciencedatabaseswereincluded,such

asScienceDirect,inordertobroadenthesearchresultstoensuregreatercoverageofthe

entireliteratureintothesecondarytransferofDNA.

34

Thekeytermsusedforsearchingthedatabasesincludedthefollowing:

• “Secondarytransfer”AND“DNA”AND“Forensics”

• “DNAtransferevents”AND“Forensics”

• “Tertiarytransfer”AND“DNA”AND“Forensics”

• “DirectDNAtransfer”AND“Forensics”

• “Shedderstatus”AND“Forensics”

• “Indirecttransfer”AND“DNA”AND“Forensics”

Noadditionalsearchtermswereexcludedduetothespecificityofthesedatabasesand

theirsmallerrangeofjournalarticlesavailable.

Theonlyrestrictionsthatweremadeonthedatabasesearcheswerethatofeliminating

types of articles, e.g. book chapters, encyclopedias etc.No additional restrictionswere

placedonthedatabasesearches.Alldatabasesalreadyhadrestrictionssuchaslanguage

anddatesappliedtosearches,whereEnglishwasselectedasthelanguageandbeginning

datesofsearcheswenttothebeginningofthedatabase.Thisdatewasmostcommonlyin

the1990’sandsearchresultswenttoApril2018.

3.1.1SCREENINGOFARTICLES

After thecompletionof thedatabasesearches,allarticleswerethenscreenedfor their

appropriatenesstothetopicofthesecondarytransferofDNA.Atotalof307articleswere

identifiedthroughthedatabasesearch.Thesearticleswerecompiledofallyieldedsearch

resultsfromallsixsearchtermsthroughallsixdatabases.These307articlesthenhadtheir

reference lists screened yielding a further 103 eligible articles, creating a total of 410

articlesthroughtheliteraturesearch.

35

Thenextstepoftheprocesswastoscreenall410articlesfoundasaresultoftheliterature

searchbasedontheirtitlesandabstracts.Forthis, inclusionandexclusioncriteriawere

usedtonarrowdownthepossiblearticles.Themainexclusioncriteriathatwasusedwasif

thearticledidnotinvolvetransfereventofDNAinanyform(biologicalsubstancesuchas

touch,spermatozoa,bloodandsaliva)itwouldbeexcludedfromthemeta-analysis.Other

exclusioncriteriaincludedstudiesthatinvolvesacasestudy,thosethatarearesponseto

ajournalarticleandaformofwritingthatwasnotajournalarticleortheses(e.g.bookor

abookchapter).Fromthisscreeningprocess,atotalof141articleswereselectedbased

ontheirtitleandabstracts.

These 141 articles then went through a further screening process that involved the

inclusionoftheentirebodyoftext.Articleswereagainexcludedbasedoninclusionand

exclusion criteria, where if an article had a recorded or measured occurrence of the

secondarytransferofDNAinanyform,itwasabletobeencompassedinthemeta-analysis.

ArticlesthatalsofoundanoccurrenceofthesecondarytransferofDNAbutindirectly(was

notthemainvariabletestingfor)werealsoexcluded.Thesecondscreeningresultedina

total of 38 articles that were able to be continued through to themeta-analysis. This

resultedin103studiesexcludedfromthisscreeningprocess.

3.1.2DATAEXTRACTION

Data was extracted from all 38 articles discovered through the screening process and

collatedintoatableusingMicrosoftExcel.Thisdataincludedthefollowing:

• Author;

• Year;

36

• Conditionsoftheexperiment(Controlled,semi-controlled,uncontrolled);

• NumberofParticipants;

• Numberofsamplestaken;

• Typeofvector;

• FinalobjectoftheDNAdeposition;

• Timebetweentheprimaryandsecondarytransferevent;

• Timehandlingthevector;

• Timehandlingthefinalobject;

• Typeandpressureofcontactofvector;

• AveragetotalDNArecovered(ng);

• TotalDNAconcentration(ng/µL);

• Totalnumberofunknownallelesdetected(indicativeofsecondarytransfer);

• Numberofmixturesseen;

• Ifareferencesampleofvolunteershadbeentaken;and

• IfsecondarytransferofDNAinanyformwasdetected.

3.2RESULTSANDDISCUSSION

Itwasoriginallyhypothesizedthatthroughtheexplorationofliteratureandcompletionof

ameta-analysisthatanappropriateandconciseguidewillbeproducedtodeterminethe

chancesofasecondarytransfereventoccurringandwhatconditionsarerequired,toaid

Biologists inexperttestimony.Thishypothesiswasrejectedonthebasisthatthemeta-

analysiswasfoundnotbeanadequatemodelforthedatathatwasavailable.Thescreening

processofarticlesexploringthesecondarytransferofDNAwassufficientforthisfieldof

37

science,asarticleswereabletobeeasilyincludedandexcludedbasedonasetofcriteria.

Dataextractionwasadditionally able tobe carriedout to someextent asmost journal

articlesgiveaccuraterepresentationsofthestudyandhowitwasperformed.However,

through the creation of a table, incorporating all of the included studies for themeta-

analysisandtheconductionofthemeta-analysis,itwasdeterminedthatthemeta-analysis

wouldnotbeadequateforthedatathatwasavailable.

Forthismeta-analysis,itwashopedtouseaverageamountoftotalDNArecovered(ng)as

oneofthequantitativevariables.Articleshoweverdidnotallgiveresultsthatwouldbe

sufficienttocalculateanaveragefrom.SomegaveallvaluesoftotalDNArecoveredfrom

thefinalobjectsaftersecondaryDNAtransfer,whereanaveragewasabletobecalculated.

Conversely,somearticleswouldstatearangeofthetotalamountofDNArecoveredwhich

anaveragecouldnotbecalculatedfromastherawresultswerenotpresent.Therefore,

therewasnotaconsistentsetofvaluesthatwouldbeusedtorepresentthetotalamount

ofDNArecoveredthroughallincludedstudies.

Asasecondquantitativevariablewasrequired,outofthedataavailabletoextract,time

handlingthevector,finalobjectortimebetweenhandlingthevectorandfinalobject,were

theonlytwooptionsavailable.Althoughthisvaluedifferedbetweenstudies,withinastudy

itwashighlylikelythatonlyasingletimevaluewasutilized.Forexample,Gorayetal.used

oneminuteforthetimehandlingthevectorandthefinalobjectforallvolunteers.27Two

different vector types were used within the study with two different times between

handlingthevectorandfinalobject(immediatelyand24hoursafter).27BothtouchDNA

and bloodwas used as the biological substances, creating four possible scenarios.27 As

38

thesequantitativevariablesdidnotvary substantiallywithin thestudy, itwouldnotbe

possibletocomparethevariableswithinthestudy.Otherstudiesadditionallyonlyhadone

vector typeandbiological substance,andthereforeonlyonequantitativevalueof time

handlingvectorsorobjects.28,29,35,41,45

Reflecting back onto the hypothesis, it was stated that it was anticipated that “an

appropriateandconciseguidewillbeproducedtodeterminethechancesofasecondary

transfereventoccurringandwhatconditionsarerequired”.Asmentionedpreviously,the

frameworkthatameta-analysisworksonisthecomparisonoftwoquantitativevariables

inthehopetoperformstatisticaltestssuchasCohen’sDorPearson’scoefficient.Inorder

to determine the conditions required for the secondary transfer of DNA to occur,

quantitativevalueswouldneedtobegathered.TheconditionthatsecondaryDNAtransfer

are conducted under would include things such as the vector type, final object type,

conditionsoftheexperiment,backgroundDNApresentandtypeofpressureandcontact

ofthevectororfinalobject.Thesevariablesunfortunatelyarequalitativeanddonothave

an option to measure them quantitatively. In that case, performance of comparisons

betweenthesevariablesandquantitativevariablessuchastotalDNA,DNAconcentration

ortimehandlingobjectscannotbecarriedout.

4. LIMITATIONSOFTHESECONDARYTRANFEROFDNA

4.1CONTROLLEDNATUREOFTHESTUDIES

Withavastnumberofpapersexploredinthisliteraturereview,noneofthemtestedthe

occurrenceofsecondaryDNAtransferhappeningwithcompletelyuncontrolledscenarios.

39

In my opinion, the closest study that was to an uncontrolled scenario was a study

conductedbyGoray&vanOorschotin2013.Theyusedasmallgroupofindividualswitha

smallnumberofobjects(glasses,jugs,tables,chairs)participatinginsocialinteractionfor

20minutes.31Priortotheexperimentoccurringtheauthorsdidpre-cleanallsurfacesand

itemsandthemajorityofcontrolsproducedanegativeresultasexpected.Ascloseasthis

studywastobeinguncontrolled,theoccurrenceofpre-cleaningdiminishesallbackground

DNApresentonthoseitems,notreflectingwhatweretooccurina“real-life”situation.

4.2TIMEBETWEENTAKINGSAMPLES

Thereseemstobeacommonthemebetweenstudiespublishedthatmostsamplesfrom

transfereventsaretakenalmostimmediatelyaftertheeventoccurs.Asstatedinoneof

thestudies,“thebestchanceofrecoveringtheDNAwouldbetosampleassoonaspossible

afterthefinaltransfereventhasoccurred”.41Thisunfortunatelyisnotpossibleinreal-life

situations as often a crime scenemay not be discovered for hours or days after it has

occurred. This same principle can be applied for the time between transfer events

occurring.Loweetal.hadfoundwhenplacinga30seconddelaybetweentransferevents

the final sample went from containing a full profile with all the first individuals’ DNA

presenttoonlyhave70%ofthatsameindividualspresent.42Evenwiththat30secondtime

delay,thesituationisstillnotindicativeofreal-lifeasindividualswouldnotshakehands

immediatelypriortocommittingacrime.Judgingofftheseresults, itcouldbesaidthat

unlessthesesituationsweretohaveoccurredandtheadditionofsomeothervariablesit

wouldbehighlyunlikelythatDNAfoundatacrimescenewouldbeanyoneelse’sother

thanthatindividualcarryingoutaprimarytransfer.

40

4.3REFERENCESAMPLES

Innearlyallofthestudiesexploredthroughoutthisreview,referencesamplesweretaken

from thevolunteers.This couldbeconsiderednormal for studies in this field,although

whentryingtobestrepresentwhatoccursinthereal-world,thisactiondoesnotcorrectly

reflect that.When interpretingmixturesor just complex single sourceprofiles, forensic

scientistswillmorelikelythannotneverhaveareferencesampleforcomparisonpurposes.

Hereitmaythenbemoredifficulttopulloutmultiplecontributorsofamixturewithout

knowledgeofthemajorcontributor,oranycontributor.Byusingareferencesampleofthe

volunteerduringthesestudies,essentiallymostoftheworkforensicscientistsconductin

regardtoprofileinterpretationisvoid.

5. LIMITATIONSOFMETA-ANALYSIS

5.1NEWAPPLICATIONTOTHISFIELD

Meta-analysesareaverycommonmethodappliedtothatofthemedicalfieldcurrently

andaspreviouslymentioned,ithasnotyetpreviouslybeenappliedtothefieldofForensic

Science.Becauseofthisitwasnotknownifthemethodcouldbeappliedinthesameway

asithasdonebeforeinthemedicalfield.Asdiscoveredthroughattemptingameta-analysis

on thesecondary transferofDNA, itwasnotanappropriatemethodon these typesof

studiespublished.

5.2TYPEOFRESULTSPUBLISHED

Throughconductingameta-analysisofthesecondarytransferofDNA,itwasdetermined

thatthestudiesconductedandpublishedwerenot inaformthatdataextractioncould

41

occurefficiently.Notallstudiespublishedtheirresultsinthesamewayanditwasdifficult

to obtain two quantitative variables for comparison. Close to none of the 38 studies

publishedinthesecondarytransferofDNAandincludedinthemeta-analysis,published

theirrawresultsofvariablessuchastotalamountofDNArecovered.Duetothis,averages,

meansorstandarddeviationswerenotabletobecomputedforresultsincludedinthese

studies,andthereforeameta-analysiswasnotabletobecarriedout.

6. THEFUTUREFORTHESECONDARYTRANSFEROFDNA

Themainpointthatshouldbetakenoutofthisreviewisthatinthisfieldweneedmore

studies thatexamine theoccurrenceof the secondary transferofDNA inmore life-like

scenarios.Untilthen,onlysomuchcanbespeculatedaboutifthistransfereventoccurs

enough to be a valid argument in court for why an individuals’ DNAmay be present.

Although performing these uncontrolled studies may not produce high instances of

secondaryDNAtransferoccurring,throughanapplicationofameta-analysisresultsfrom

eachstudycanbecombinedtoproduceahigherstatisticalpower.

7. CONCLUSION

Atthisstage,itcanbeconcludedthatthehypothesis,throughtheexplorationofliterature

andcompletionofameta-analysisthatanappropriateandconciseguidewillbeproduced

todeterminethechancesofasecondarytransfereventoccurringandwhatconditionsare

required,toaidBiologistsinexperttestimony,canberejected.Datapublishedinthe38

studiesincludedforthemeta-analysiswasnotadequateenoughtoperformquantitative

comparisonsbetweenvariables inorder tocombineresults tostrengthenthepowerof

42

eachstudy.Infuture,itshouldbeconsideredthatarticlespublishedinthefieldofForensic

Sciencedisplaytheirresultsindifferentwaysorgiveaccesstoreadersoftheirrawresults

forfurtherinterpretationpurposes.

43

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THISPAGEHASBEENLEFTINTENTIONALLYBLANK

49

PARTTWO

MANUSCRIPT


50

ABSTRACT


bodythatcontainsallofanindividual’sgeneticmaterial.TraceDNAhasallowedDNAtobe

detectedoneverydayobjectssuchasdoorhandles,tablesandchairs.Currently,theidea

ofthesecondarytransferofDNAhasarisen,involvingthetransferofgeneticprofilesfrom

anindividualthroughavectorandthentofinalindividualorobject.Thishasbecomeare-

occurringterminthecourtsystem,asindividualsstandingtrialusethisargumentforthe

presence of their DNA. It is therefore hypothesized that through the exploration of



conditions are required, to aid Biologists in expert testimony. Database searcheswere

conductedtogainresourcesinthetopic,followedbyanumberofscreenstodetermine

the suitability of articles for the meta-analysis. With the resultant 38 articles, data

extractionwasadditionallycarried.However,duetothelackofquantitativevariablesand

results,ameta-analysiswasnotabletobeconducted.It issuggestedthatinthefuture,

studiespublishtheirresultsinamorescientificallyrigorousmannerorallowtheaccessof

rawresultsforexternalinterpretationpurposes.

Keywords:forensicscience,secondarytransfer,DNAtransfer,indirecttransfer,touchDNA,

DNA

51

TABLEOFCONTENTS

ABSTRACT………………………………………………………………………………………………....................50LISTOFABBREVIATIONS………………………………………………………………………………...............52INTRODUCTION………………………………………………………………………………............................53MATERIALSANDMETHODS………………………………………………………………..........................55

DATABASESEARCHES……………………………………………………………...............................55SCREENINGOFARTICLES………………………………………………………...............................56DATAEXTRACTION………………………………………………………………................................57

RESULTSANDDISCUSSION……………………………………….....…............................................58LITERATUREREVIEW……………………………………………......…….....................................58

OBJECTASVECTOR………………………………………………....................................58 INDIVIDUALASVECTOR…………………………………………...................................61 INDIRECTFINDINGSOFSECONDARYTRANSFER……………...........................62 SHEDDERSTATUS…………………………………………………….................................64 OTHERFORMSOFDNAINVOLVEDINSECONDARYTRANSFER…………….....65 TERTIARYTRANSFEROFDNAANDBEYOND…………………………....................67 CASESTUDIES………………………………….........……........………….........................69 LIMITATIONS……………………………………………………………................................69META-ANALYSIS…………………………………………………......……...........……………..............71

CONCLUSION……………………………………………………………………………………………..................74REFERENCES…………………………………………………………………………………………......................75

52

LISTOFABBREVIATIONS

DNA=DeoxyribonucleicAcid

ng=Nanograms

ng/µL=Nanogramspermicroliter(concentration)

pg=Pictograms

STR=Shorttandemrepeat

µL=Microliter

UV=Ultraviolet

53

INTRODUCTION

Deoxyribonucleicacid(DNA)ismoleculepresent innearlyeverycellofthehumanbody

thatpossessallofanindividual’sgeneticmaterial.1Eachcellhasapproximatelythreebillion

basepairswhichisessentialforthecharacterizationofeachindividual.1Thesebasepairs

cancreatepolymorphicsequences,forexampleashorttandemrepeat(STR),arepeated

sequenceofDNAthatisusuallyaroundonetosixbasepairsinlength.2Theseareasareof

greatinterestinthefieldofforensicscience,astheircombinationsarevariablebetween

individuals.InForensicScience,STRtypingisusedtoinferthenumberofrepeatsofDNA

where today kits such as PowerPlex 21® and GlobalFiler® are used within forensic

laboratoriesinAustralia.

TraceDNAhasbecomemoreestablishedoverthepastcoupleofdecadesinthefieldof

forensic science. It is defined as cells off the skin surface that have been carried onto

anothersurfacethroughphysicalcontact,allowingdetectionofDNAoneverydayobjects.3

TraceDNAwasfirstexploredingreatdepthbyvanOorschotandJonesin1997,wherethey

found that they could obtain DNA profiles from swabbing objects that were regularly

handledbyaparticular individual.4Previouslyapproximately1ngofDNAwasneededto

gainafullprofilebutnowapproximatelyonly100pgisneeded.5,6Duetothisincreasein

sensitivityofthetechnology,conceptssuchassecondaryandtertiarytransferofDNAhave

nowdeveloped.

ThesecondarytransferofDNAisdefinedasthetransferofDNAfromonepersonorobject

toanotherviaan intermediatepersonorobject,not involvingadirect linkbetweenan

individualandthetargetsurface.6-9Withcontinuedresearchcanresult inanincreaseof

54

knowledge,wherenowconceptssuchastertiaryandquaternarytransferhasarisenthough

secondary transfer is still beingexplored. Similar to secondary transferofDNA, tertiary

transferinvolvesthetransferofDNAfromonepersonorobjecttoanotherthroughtwo

intermediary vectors.10 Quaternary transfer requires an additional transfer events,

resultinginatotaloffourtransfersofDNA.Alltransfereventshavebeenshowntooccur

innumerouspublishedpapers,butnotconsistently.11-15

Since itsfirstuse inthecourtsystemsin1986,DNAhasbeenahighlyregardeditemof

evidence, something that has been very hard to argue against. AsDNAbecomesmore

understood,scientistsareabletogainagreater insight intowhohasbeenpresentata

crimesceneandincreasedsensitivityofkitsallowsforensicscientiststogainDNAprofiles

fromthesmallestofbiologicalsubstances.ClaimsofthesecondarytransferofDNAcan

nowoccurduetothis increasedknowledgeofDNAandsensitivityofkits,meaningthat

nowanindividualmaynotdirectlydeposittheirDNAontoanobject,butrathertransferred

throughavector.This isbecoming increasingly commonwithin thecourt systemanda

need for a concise guide to determine the possibility of secondary transfer has now

emerged.

Aproposedsolutionisthroughameta-analysis,ananalysisthatpoolsresultsfrommultiple

studiestogethertoincreaseitsstrengthandsamplesize,thusthepowerofthestudies.3,16

Tocarryoutacorrectmeta-analysis,itshouldbedoneasapartofasystematicreviewand

includefeaturessuchasacomprehensive,reproduciblesearchandtransparentselection

criteria.16Therearetwomaintypesofreviews,systematicandnarrative.Narrativereviews

exploremorebroader topicsandqualitatively summariseevidence,whereassystematic

55

reviewsfocusmoredeeplyonprimarystudieswhoexploreclinicalquestions.16Thereare

twotypesofmodels thatareusedto forameta-analysis, randomeffector fixedeffect

models.16 The fixed effect model assumes that the ‘true’ effects are fixed whereas a

randomeffectmodelassumesthatonlyarandomsampleisincluded.16Therandomeffect

modelismorepreferredforameta-analysisasitbetterreflectsrealityasstudiesbarely

evermimiceachother.16Thisisthereforewhyarandomeffectmodelwillbeusedforthis

meta-analysis.

Theaimofthis literaturereviewwastoproduceacombinedreviewintothesecondary

transferofDNAinadditiontodrawingmultiplestudiestogethertoattempttoprovidean

analysis of the literature. It is therefore hypothesized that through the exploration of



conditionsarerequired,toaidBiologistsinexperttestimony.

MATERIALSANDMETHODS

DATABASESEARCHES

Aliteraturereviewwasconductedinconjunctionwiththemeta-analysiswhereresources

weregatheredusingonlinedatabasesintothesecondarytransferofDNA.Databaseswere

chosenbasedontheirrelevancetothefieldofForensicScience.Chosendatabasesincluded

Scopus,WebofScienceCoreCollection,ProQuest,Medline,ScienceDirectandResearch

Gate.Theseincludedbothspecificandgeneraldatabasestoensurehighcoverageofall

56

published literature inthesecondarytransferofDNA.Keytermsusedforsearchingthe

databasesincludedthefollowing:

• “Secondarytransfer”AND“DNA”AND“Forensics”;

• “DNAtransferevents”AND“Forensics”;

• “Tertiarytransfer”AND“DNA”AND“Forensics”;

• “DirectDNAtransfer”AND“Forensics”;

• “Shedderstatus”AND“Forensics”and

• “Indirecttransfer”AND“DNA”AND“Forensics”.

Noadditionalsearchtermrestrictionswereappliedduetothespecificityofthedatabases

andthe lackof functionprovidedtodoso.Theonly restrictionthatwasappliedto the

database searches were those that eliminated specific types of resources, e.g. book

chapters&encyclopedias.Alldatabasesalreadyhadrestrictionsappliedtosearchessuch

aslanguageanddates,whereEnglishwasthelanguageanddatesrangedfromthe1990’s

toApril2018.

SCREENINGOFARTICLES

Atotalof307articleswereidentifiedthroughthedatabasesearch,compiledofallyielded

searchresults.Allarticlesthenhadtheirreferencelistsscreenedresultinginafurther103

eligiblearticles,creatingatotalof410articlesidentifiedthroughtheliteraturesearch.The

nextstepwastoscreenall410articlesusingtheirtitlesandabstracts,basedoninclusion

andexclusioncriteria.Themainexclusioncriteriathatwasappliedwasifthearticledidnot

involvethetransfereventofDNAinanyform(e.g.touchDNA,spermatozoa,bloodand

saliva) it would be excluded from the meta-analysis. Other exclusion criteria included

papersthatinvolvesacasestudy,thosethatarearesponsetoajournalarticleandaform

57

ofwritingthatwasnotajournalarticleortheses(e.g.bookorabookchapter).Fromthis

screeningprocess,atotalof141articlesweredeemedsuitable.These141articlesfurther

had theentirebodyof text screened, againusing inclusionandexclusion criteria. If an

articlehadarecordedormeasuredoccurrenceofthesecondarytransferofDNAinany

form, it was able to be encompassed in themeta-analysis. Articles that discovered an

occurrence of the secondary transfer ofDNAbut indirectly (was not themain variable

testingfor)werealsoexcluded.Thisscreenresultedinatotalof38articlesthatwereable

tobecontinuedthroughtothemeta-analysis,and103studiesexcluded.

DATAEXTRACTION

Data was extracted from all 38 articles discovered through the screening process and

collatedintoatableusingMicrosoftExcel.Thisdataincludedthefollowing:

• Author;

• Year;

• Conditionsoftheexperiment(Controlled,semi-controlled,uncontrolled);

• NumberofParticipants;

• Numberofsamplestaken;

• Typeofvector;

• FinalobjectoftheDNAdeposition;

• Timebetweentheprimaryandsecondarytransferevent;

• Timehandlingthevector;

• Timehandlingthefinalobject;

• Typeandpressureofcontactofvector;

58

• AveragetotalDNArecovered(ng);

• TotalDNAconcentration(ng/µL);

• Totalnumberofunknownallelesdetected(indicativeofsecondarytransfer);

• Numberofmixturesseen;

• Ifareferencesampleofvolunteershadbeentaken;and

• IfsecondarytransferofDNAinanyformwasdetected.

RESULTSANDDISUCSSION

LITERATUREREVIEW

RegardinganytransfereventofDNA,thewayinwhichitoccursisrelativelysimilaracross

theboard.TheremustalwaysbeaprimarytransfereventinvolvingthemovementofDNA

from an individual to either an object or another individual. There then must be a

subsequenttransferevent,wherethatsamegeneticprofilewillbetransferredeithertoan

individual or an object. In this case, it could be hypothesized that the more common

scenariowouldbethatofanindividualasthevector.Belowstudiesusingbothinstances

asvectorswillbeexploredandreviewedtogainagreaterunderstandingastohowthis

mechanismoccursanditslikelihood.

OBJECTASVECTOR

When the secondary transfer of DNA is put forth as a possible explanation, it ismore

sensibletohaveoccurredthroughanobjectratherthananindividual.Oneofthefirststudy

thatperformedsecondaryDNAtransferthroughanobjectwasperformedin2012.Thefirst

59

experimentsawhandscomeincontactwithatoyorasingletandthenthatobjectcoming

incontactwithalaboratorycoat.17Resultsshowedthatin19outof20repeats,DNAfrom

thedonorwasobserved.17AuniqueapproachwasusedbyFrenchetal.whereUltraviolet

(UV)powderwasusedthroughthetransferexperiment.Threeparticipantssatarounda

tabledrinkingfromabottlewiththeirownthreeglassesfor30minuteswhereonlyone

individualstartedwiththeUVpowder.18Therewasnodirectcontactbetweenindividuals

butthestudyshowedtheindirecttransferoftheUVpowdertoallindividuals.18

GorayandvanOorschotperformedaverysimilarexperimenttoFrenchetal.thefollowing

yearwherethesame“uncontrolled”conditionswereused.Herehowevertheydidnotuse

UVpowderbuttestedfortheDNAitself.In27%ofthetablesamplesandthen33%ofthe

chairsampleshadaDNAprofilesfromparticipantsthatdidnotcomeintocontactwiththe

surfaceindicatingthattheseprofilesmighthavebeentransferredfromthejug.19Afurther

58% of table samples and 42% of chair samples had DNA profiles from unknown

individuals.19

Margiottaetal.usedglovesasthevectorfortransferwhereinhalfthecasestestedfor,

they found a mixture of DNA containing alleles belonging to an unknown individual

different to that of the volunteer.20 In an additional 15%of the cases just alleles from

another unknown individual were recovered not attributed to the volunteer.20 Here

secondarytransferhasoccurred,althoughauthorsdidpointoutsomeof theirnegative

controlsfromsupposedly“clean”glovesdidyieldalleles.20Szkutaetal.furtheredworkof

otherstudiesthatfocusedonthequantityofDNAthatisleftonvectorsafteratransfer

eventhasoccurred.Secondary transfereventswereconductedbetweentwo“exhibits”

60

usingtoolssuchasforceps,scissorsandglovesasthevector.21Itwasshownthatsecondary

transfercouldoccurfromexhibittoexhibitbyDNA-freevectorsbutthemainfindingwas

thepresenceofasufficientquantityofDNAonthevectorforfurthertransferevents.21

Fonneløp et al. investigated the transfer of DNA based on a case study, using three

volunteerswhowereknowntobegoodshedders.12Thefirstindividualhandledanobject

for 30 seconds followed by a second individual.12 Results showed for the first transfer

event,83%oftheDNAwashighenoughqualityforprofiledetectionandforthesecond

transfer event, that figure dropped to 53%.12 In 2017, Buckingham et al. furthered a

previousstudywhereporouswereusedratherthannon-poroussurfacestoinvestigatethe

resulting yield and profile of DNA. Some results produced evidence of secondary and

tertiary transfer ofDNA although the proportion to the depositor’sDNAwas less than

10%.22

Theuseofalaundrymachineforthetransferofspermatozoaisarelativelywellcovered

topic.AstudyconductedbyKamphausenetal.in2015wasoneofthefirsttestingforthe

transferoftouchDNAhoweverthesecondarytransferoftouchDNAwasdeemedalmost

impossibleforreliableSTRanalysis.23Asecondstudyconductedin2018sawunwornsocks

werewashed inanormal loadofhousehold laundry.1522%of samplesyieldeda result

higher than theminimal threshold forcasework (0.06ng/µL).15Foursamplesmatcheda

profile of a female from thehousehold indicative of the transfer ofDNAoccurringbut

unknownastoeitherdirectorindirecttransfer.15

61

VanOorschotetal.wasthefirsttotesttheextenttowhichDNAtransfermayoccurusing

fingerprintbrushes.2473currently-usedfingerprintbrushedwerebrushedover5sheetsof

plastic, includingonecontaininga freshdepositedhandprint.24Theresultsshowedthat

verylimitedpickupandtransferofDNAoccurredbythebrusheshoweverwhenchanges

weremadetotheDNAanalysis,significantpickupandtransferofDNAwasseen.24Asecond

studywasconductedbyProffetal.wheretheydeterminedthatDNAcontaminationwith

fingerprint brushes was quite common (85%) and for the secondary transfer, 25% of

surfaceshadDNAdetectedwithonefullprofile.25Aswiththepreviousstudy,theyalso

foundalimitedriskofthesecondarytransferofDNAthroughfingerprintbrushes.24,25

INDIVIDUALASVECTOR

This scenario appears to be themore common scenario to occur although is the least

researched.ThefirststudythatusedanindividualasavectorforthetransferofDNAwas

conductedby Loweet al. in2003. The study found thatwhenpoor shedderswere the

vector,more often than not, the original individual had their full profile transferred.26

Additionally,witha30secondtimedelaybetweentransferevents,mixedprofileswere

discovered where 70% were from the original source.26 Farmen et al. additionally

conductedastudywheretwoindividualsundertookahandshakefollowedbythegripping

ofbeakers.9Althoughnotspecified, theauthorssuggest thatall samplesweremixtures

containingbothprofilesoftheknowncontributors,indicatingthatsecondaryDNAtransfer

hadsuccessfullyoccurred.9

AstudyconductedbyMeakinetal in2015alsoshowedthesecondary transferofDNA

throughahandshakeandthenhandlingaknife.Thestudyfoundthatthreeofthefoursets

62

ofknives,mixtureswerefoundcontainingthreedifferentprofiles.27Twoprofilescouldbe

attributedtothetwovolunteers,buttheotherwasanunknownprofile,possiblythecause

of secondary or tertiary transfer.27 Cale et al. performed a standard study into the

secondarytransferofDNAwithanewkitwithincreasedsensitivityduetoagapfoundin

theliterature.TheauthorsaimedtostudyhowthepresenceofsecondaryDNAtransfer

wouldaffecttheinterpretationofDNAtypingresults.6Dataobtainedfromfiveofthe24

samples suggest that individuals can have their DNA deposited on objects in sufficient

quantitiesthattheycanbeidentifiedastheonlyormajorcontributor,suggestingthatthey

hadcomeindirectcontactwiththeobject.6

INDIRECTFINDINGSOFSECONDARYTRANSFER

TherearealsoinstancesthatoccurinwhereprimarytransferorthepersistenceofDNA

thathavebeenexploredduringastudyalthoughunknownallelesaredetected.Here,these

studies thathavedetected secondary transferofDNAduring another experimenthave

beencollated.However,inthesestudies,unlesstheauthorshavespecificallystatedwhere

thesecondarytransferhasoccurredandwhotheunknownallelesbelongto,wedonot

knowtheoutcome.Additionally,unknownallelesattributedtobackgroundDNApresent

onaregularly-usedobject.

ThefirstinstanceofaccidentaloccurrenceofsecondaryDNAtransferwasduringastudy

conductedbyLoweetal.in2002wheretheauthorstestedshedderstatusofindividuals.

Secondary transferwasdetectedwhenparticipantshelda tube for10 secondsbut the

extentwasdependentonthesubjectpairings.28Afewyearslater,Phippsetal.conducted

astudyingreatsimilaritywheretheyfoundthattheoccurrenceofsecondarytransferwas

63

lowandnotlikelybutpossible.29Only14allelesoutof581couldnotbeattributedtothe

volunteersequatingto0.02%.29AstudyconductedbyRuttyfoundaninstanceofpossible

secondaryDNAtransferoccurringwheremale-femalepairswereusedinordertosimulate

astrangulationevent.30Twoswabsofthemale’sfingers(controls)yieldedaprofileofa

female inthestudyalthoughthesetwoindividualshadneverbeenpairedorcomeinto

contactwithoneanother.30

Petricevicetal.usedfivevolunteerswhosleptintheirownbedbutwithanewlowerbed

sheetor individualsslept inabedwheretheyneverhadpreviously.31When individuals

sleptintheirownbeds,asecondindividual’sDNAprofilewasfoundinatleastonesample

from3outofthe5volunteers.31Whenindividualssleptinanunknownbed,allunknown

samplescouldbeattributedtotheregularuser’sprofilewhichdidnotindicatesecondary

DNAtransfer.Djuricetal.hadvolunteersholdplastictubesandindividual’sanklesfor10

seconds.32Whenvolunteersheldtubes,outofthesevenprofilesobtains,oneresultedin

a partial profile not matching the volunteer’s, indicating the occurrence of secondary

transfer.32Whenankleswereusedasdeposition sites, nooccurrenceof the secondary

transferofDNAwasnoted.32Gorayetal.conductedaverysimilarexperimentagainbut

testedthedepositionontoglassplatesratherthanplastictubes.Atotalof40sampleswere

takenandnon-selfDNA(unexpectedalleles)wasseenin79%ofthesamples.33

ApaperpublishedbyOldonietal.appliedtheconstantunknownfromcontactstainsat

break-ins to conducted an experiment. A total of 231 samples were created where

approximately2-10%containedunknownalleles,possiblesuggestingsecondarytransfer.34

In2015,FonneløpshowedthatsecondarytransferofDNAoccurredthroughthepresence

64

oftheoriginaluser’sDNAonthehandsoftheseconduserupto8daysafterreceivingthe

computerequipment.35Lacerenzaetal.directlyswabbedindividual’shand’swhereoutof

120samples,56hadoneormoreunknownallelesand36ofthosecouldbeclassifiedas

mixtures(64.3%).37InastudyconductedbyvandenBergeetal.,individualsdraggedeach

othermimickinganactivity-relatedscenario.Outof26samples,3ofwhichdidnotresult

inover7alleles.37Donoralleleswerepresent in100%ofthesamplesandnon-donorin

71%.36Morerecently,Mageeetal.usedcollarsandcuffsofuppergarmentstodetermine

the presence of DNA.38 Out of 55 samples taken, non-wearer DNA was recovered, an

averageof1.3ngfromthecollarand2.7ngfromthecuffs.38

SHEDDERSTATUS

Shedderstatusisoneofthemorerecentconceptsundertheumbrellaofthetransferof

DNAandonethatwillcontinuetogrowasmoreresearchisconductedintothetopic.The

first study to directly carry out an experiment on individual’s shedder status’ was

conductedbyLoweetal.in2002andisconsideredthebackbonestudytothistopic.The

authorsgotparticipantstogripasteriletube,15minutes,2hoursand6hourperiodsafter

hand-washing.29 They determined that there were differences between individuals in

regardtohowmanyallelesthattheydepositedonasurface.29Goodshedderswerethen

classediftheindividualleftanentireprofileafter15minutesofhand-washing.29Allother

individualswereclassedaspoorshedders.

Phippsetal.conductedasecondstudythatinvestigatedshedderstatusandusedalmost

identical experiments to that of Lowe et al. Unexpectedly, the study had shown that

individualsdonotproduceconsistentquantitiesofDNAovertime,varyingasmuchwithin

65

themselvesthancomparedtootherpeople.30UsingthecriterialaidoutinLoweetal.,this

studydiscoveredno“good”sheddersandallwereclassifiedas“poor”shedderssuggesting

thislevelofclassificationmaynotbeallassimpleasthought.29,30Farmenetal.measured

individuals’ shedder status just based on swabs from their hands.39 It did not test the

quantityofDNAthattheindividual’sactuallydepositonasurface,soitmaynotclassasa

truerepresentationofshedderstatus.Outofthenineindividualswhoparticipatedinthe

study,twoindividualshada lowDNAyieldandsevenindividualsproducedprofilesthat

were able to be analysed.39 More recently, Goray et al. collected 240 samples from

handprintsonglassplateswhereonlyfourofthesampleshadnodetectablequantitiesof

DNA.33Twoofthetenparticipantscouldbeclassedasgoodsheddersbutthiswasdone

underdifferentcriteriaandexperimentalcriteriaastothebasestudyconductedbyLowe

etal.33

OTHERFORMSOFDNAINVOLVEDINSECONDARYTRANSFER

Oneof the first studiescarriedout in thebroader realmof the transferofDNAwasby

Kafarowskietal.in1996.Conductedbeforethefirstoccurrenceofthesecondarytransfer

oftouchDNA,itaimedtodeterminethelikelihoodofthetransferofspermatozoabetween

articlesofclothingduringamachine-cyclewash.40Althoughtheretentionofspermatozoa

duringmachinewashhadpreviouslybeenexplored,thetransferofittootherarticlesof

clothinghadn’t.41,42Theauthorsusedunderpantsasameansfortheprimarydepositof

spermatozoa,washingthemwiththreeadditionalcleanpairsandthenmachinedried.40In

allthreetrials,tracequantitiesofspermatozoawerefoundwhereaminimumofonesperm

headandmaximumofeightpersample.40

66

SalivaisanadditionalbiologicalsubstancethatcontainshumanDNAthathasthepossibility

tobetransferredthroughobjectsandindividualstocauseasecondarytransfer.Thefirst

thatexploredintothissecondarytransferofsalivawasconductedbyWiegandetal.where

salivawastransferredontoeitherpaper,cottonorplasticsurfaces,leftthemtoairdrythen

athumbrubbedontothestain.43Thethumbwasthenplacedontoanotherpieceofpaper

andthentheareawasswabbed.43Intotalonly1outof96gavecompleteprofiles,allfrom

plasticastheoriginalsource.43Vectorsofpensandplastictubeswereusedinanadditional

study.Theirstudyconcludedthatsecondarytransferofsalivadidoccur,ingreaterlevels

of retention than touch DNA and thatmoist surfacesmay facilitate DNA transfer to a

greaterdegreethanadrysurfaceaspreviouslydiscoveredinGorayetal.2010.11,43

In2015,thetransferofsalivathroughlaunderingwasstudiedwhereindividualsdeposited

saliva onto textile cloths and then these were subsequently washedwith clean textile

cloth.23 A total of 46 independentwasheswere completed both by hand andmachine

with/without detergent for saliva, blood and touchDNA.23 Salivawas seen to undergo

transfereventswhichsupportsotherstudiesperformed.11,23,24,43,44Themostrecentstudy

conducted was using individual’s hands as vectors for the transfer. 14 areas of the

individuals palm were swabbed in addition to the glass plate which the saliva was

transferredto.45Thestudyyieldedanumberofunknownallelesonthefinalplatewhere

theauthorscontributedthis toprior interactionwith individuals,supportingthefinding

thatsecondarytransferofDNAhadoccurred.45

Wiegandetal.conductedastudyintotheriskofthetransferofbloodstainsfromsurfaces

suchasplastic,cottonandpaper.Thebloodstainswereairdriedthenanindividualplaced

67

theirthumbonthestainthenontoanotherpieceofpaper.43SecondarytransferofDNA

wasfoundtooccuronthecottonsurface,bothwithandwithoutgloves.43Lehmannetal.

conducted a very similar experiment but all transfer events were conducted with

substratesonlyandnohumaninteraction.Theexperimentsupportedtheexistenceofthe

secondarytransferofDNAthroughbloodandadditionallyfoundthatifthebloodwerewet,

itwouldtransferfurtherandwouldendinahigherconcentrationasmoresubstancewas

transferred.46

Kamphausen et al. investigated the secondary transfer ofDNA through laundering in a

washingmachineorbyhand.Artificialbloodstainswerecreatedontextilecloth,driedand

thenwashedwithcleantextilecloths.23Through46independentwashes,transferofblood

cellswasseensupportingpreviousstudies.23,43,46,47Agaphadthenbeenidentifiedinthe

literature,wherenoneofthepreviousstudieshadthoroughlytestedtheeffectofaridity

ofblood.vanOorschotetal.useddryingtime,temperatureandhumidityasdependent

variablesandcottonas the secondary substrate.47The secondary transferofbloodwas

seen to have occurred with significant differences in the transfer percentage of blood

between5and60minutes.47

TERTIARYTRANSFEROFDNAANDBEYOND

Although the secondary transfer of DNA is still in the process of gaining a greater

understandingofitsmechanisms,throughthesestudiesthediscoveryoftransferevents

occurring after a second transfer have occurred. These studies are still in their very

preliminarystagesandlikesecondaryDNAtransfer,stillrequireagreaterunderstanding

intothemechanismssurroundingtheprocess.Thefirststudythatexploredtheoccurrence

68

ofthetertiarytransferofDNAwasconductedbyWarshaueretal.whereanobjectwasthe

original vector andwas further transferred from an individual to another individual or

objectasthetertiarytransferevent.11TheywereabletoprovethattertiarytransferofDNA

couldoccur,where87.5%displayedlessthanhalfoftheexpectedalleles.11Theseresults

correlatedwiththoseproducedbyFonneløpetal.whereafterthefinaltransfersteponly

17% of samples were high quality that could be reported and go through database

searching(5outof30transferevents).12

In the study that was conducted by Fonneløp et al. they also tested the instance of

quaternarytransferoccurring.Theyfoundthatonceagain,itoccurredwhereatotalof17

out of the 108 analysed samples contain unknown alleles.12 In 2016, Helmus et al.

performedthefirststudysolelytestingthetertiarytransferofDNA,agapidentifiedwithin

theliterature.Outof180samplesgenerated,only72haddetectableDNAthatidentified

thattertiarytransferhadoccurred(40%).12Inastudycompletedtheyearearliertheyhad

additionallyshowntheoccurrenceofthetertiarytransferofDNAbuttheauthorsnoted

thattheywereunabletodistinguishbetweentertiarytransferorinterferencefromalow-

levelofnon-donorDNA.13Inamorerecentstudy, tertiary transferofDNAevolvedtoa

morecomplexmechanismthanthatofindividual-objectinvolvedtransfer.Sixnewunworn

socksandat-shirtwerelaunderedtogetherwithnoadditionalitemsinamachinethathad

beenpreviouslyused.15ResultsshowedthattherewerenoDNAresultsrecoveredfrom

quantity (above threshold) through to profiling, suggesting that tertiary transfer didn’t

occur.15

69

CASESTUDIES

In 2017, an article was published following the incidence of contamination by Police

Officer’s in Austriawhichwithout elimination databases,many instances of indirect or

secondarytransferofDNAmaygoundetected.48Duringtheperiodof2000to2016,atotal

of 46,000 trace sampleswere submitted throughDNA and out of those a total of 347

contamination incidents were detected through the use of the elimination database

(0.75%) with a majority from indirect transfer.48 The authors further went into detail

regarding3separateincidentsthattheywereabletopinpointwherethetransferevents

hadoccurred.Thefirstincidentoccurredthroughsharedcameraequipmentthathadbeen

usedbyaPoliceOfficercompletelyunrelatedtothecase.48Thesecondshowstheindirect

transferbyaPoliceOfficerthroughasharedvehicletoavolumecrimecaseandthethird

throughpackagingofevidencematerialonanunrelatedPoliceOfficer’sdesk.48

LIMITATIONS

Limitations of the conduction of this literature review included the level of control of

studies,timebetweentakingsamplesandtheacquisitionofreferencesamples.Nostudies

presented in this literature reviewhad tested for the secondary transferofDNAunder

completelyuncontrolledscenarios.Thecloseststudytohavingacompletelyuncontrolled

study was conducted by Goray & van Oorschot in 2013. They used a small group of

individualswith a small number of objects (glasses, jugs, tables, chairs) participating in

socialinteractionfor20minutes.19Priortotheexperimentoccurringtheauthorsdidpre-

clean all surfaces and items. As close as this study was to being uncontrolled, the

occurrenceof pre-cleaningdiminishes all backgroundDNApresent on those items, not

reflectingwhatweretooccurina“real-life”situation.

70

Acommonthemebetweenstudiessawsamplesfromobjectsor individualsbeingtaken

almostimmediatelyafterthetransfereventoccurring.Asstatedinoneofthestudies,“the

bestchanceofrecoveringtheDNAwouldbetosampleassoonaspossibleafterthefinal

transfereventhasoccurred”.27Thisunfortunatelyisnotpossibleinreal-lifesituationsas

oftenacrimescenemaynotbediscoveredforhoursordaysafter ithasoccurred.This

sameprinciplecanbeappliedforthetimebetweentransfereventsoccurring.Loweetal.

hadfoundwhenplacinga30seconddelaybetweentransfereventsthefinalsamplewent

fromcontainingafullprofilewithallthefirstindividuals’DNApresenttoonlyhave70%of

thatsameindividualspresent.28Evenwiththat30secondtimedelay,thesituationisstill

not indicative of “real-life” as individuals would not shake hands immediately prior to

committingacrime.Basedofftheseresults,itcouldbesaidthatunlessthesesituations

weretohaveoccurredandtheadditionofsomeothervariablesitwouldbehighlyunlikely

thatDNAfoundatacrimescenewouldbeanyoneelse’sotherthanthatindividualcarrying

outaprimarytransfer.

Anothervariablewhichdoesnotmimic“real-life”situationsistheacquisitionofreference

samplesofvolunteersduringstudies.ForinterpretinganyDNAprofile,forensicscientists

willmostlikelynothaveaccesstoareferencesampleforcomparisonpurposesunlessa

known suspect had been previously profiled. Here itmay bemore difficult to pull out

multiple contributors of amixturewithout knowledgeof themajor contributor, or any

other contributor. By using a reference sample of the volunteer during these studies,

essentiallymostoftheworkforensicscientistsconductinregardtoprofileinterpretation

isvoid.

71

META-ANALYSIS

Itwasoriginallyhypothesizedthatthroughtheexplorationofliteratureandcompletionof

ameta-analysisthatanappropriateandconciseguidewillbeproducedtodeterminethe

chancesofasecondarytransfereventoccurringandwhatconditionsarerequired,toaid

Biologists inexperttestimony.Thishypothesiswasrejectedonthebasisthatthemeta-

analysiswasfoundnotbeanadequatemodelforthedatathatwasavailable.Thescreening

processofarticlesexploringthesecondarytransferofDNAwassufficientforthisfieldof

science,asarticleswereabletobeeasilyincludedandexcludedbasedonasetofcriteria.

Dataextractionwasadditionally able tobe carriedout to someextent asmost journal

articlesgiveaccuraterepresentationsofthestudyandhowitwasperformed.However,

through the creation of a table, incorporating all of the included studies for themeta-

analysisandtheconductionofthemeta-analysis,itwasdeterminedthatthemeta-analysis

wouldnotbeadequateforthedatathatwasavailable.

Forthismeta-analysis,itwashopedtouseaverageamountoftotalDNArecovered(ng)as

oneofthequantitativevariables.Articleshoweverdidnotallgiveresultsthatwouldbe

sufficienttocalculateanaveragefrom.SomegaveallvaluesoftotalDNArecoveredfrom

thefinalobjectsaftersecondaryDNAtransfer,whereanaveragewasabletobecalculated.

Conversely,somearticleswouldstatearangeofthetotalamountofDNArecoveredwhich

anaveragecouldnotbecalculatedfromastherawresultswerenotpresent.Therefore,

therewasnotaconsistentsetofvaluesthatwouldbeusedtorepresentthetotalamount

ofDNArecoveredthroughallincludedstudies.

72

Asasecondquantitativevariablewasrequired,outofthedataavailabletoextract,time

handlingthevector,finalobjectortimebetweenhandlingthevectorandfinalobject,were

theonlytwooptionsavailable.Althoughthisvaluedifferedbetweenstudies,withinastudy

itwashighlylikelythatonlyasingletimevaluewasutilized.Forexample,Gorayetal.used

oneminuteforthetimehandlingthevectorandthefinalobjectforallvolunteers.17Two

different vector types were used within the study with two different times between

handlingthevectorandfinalobject(immediatelyand24hoursafter).17BothtouchDNA

and bloodwas used as the biological substances, creating four possible scenarios.17 As

thesequantitativevariablesdidnotvary substantiallywithin thestudy, itwouldnotbe

possibletocomparethevariableswithinthestudy.Otherstudiesadditionallyonlyhadone

vector typeandbiological substance,andthereforeonlyonequantitativevalueof time

handlingvectorsorobjects.23,28,29,27,30

Reflecting back onto the hypothesis, it was stated that it was anticipated that “an

appropriateandconciseguidewillbeproducedtodeterminethechancesofasecondary

transfereventoccurringandwhatconditionsarerequired”.Asmentionedpreviously,the

frameworkthatameta-analysisworksonisthecomparisonoftwoquantitativevariables

inthehopetoperformstatisticaltestssuchasCohen’sDorPearson’scoefficient.Inorder

to determine the conditions required for the secondary transfer of DNA to occur,

quantitativevalueswouldneedtobegathered.TheconditionthatsecondaryDNAtransfer

are conducted under would include things such as the vector type, final object type,

conditionsoftheexperiment,backgroundDNApresentandtypeofpressureandcontact

ofthevectororfinalobject.Thesevariablesunfortunatelyarequalitativeanddonothave

an option to measure them quantitatively. In that case, performance of comparisons

73

betweenthesevariablesandquantitativevariablessuchastotalDNA,DNAconcentration

ortimehandlingobjectscannotbecarriedout.

Otherlimitationsofthemeta-analysisincludedthenewapplicationtothefieldofforensic

science,typesofresultspublishedandthesourceoftheDNA.Asmentionedpreviously,

meta-analysesaremostcommonlyusedwithinthemedicalfieldandhasnotyetbeenused

withinthefieldofForensicScience.Therefore,itwasnotknowniftheapplicationofthis

methodcouldbeappliedthesamewaythatishasdonepreviouslyinthemedicalfield.As

discovered through thispaper, itwasdeemedan inappropriateanalysis for the typeof

studiesthatarepublishedonthesecondarytransferofDNA.Thisismainlyduetothetype

of results thatarepublished.Resultswerenotpublished ina formthatdataextraction

couldoccurefficiently,additionallymakingitdifficulttoobtaintwoquantitativevariables

forcomparison.Closetononeofthe38studiespublishedinthesecondarytransferofDNA

and included in themeta-analysis,published their rawresultsofvariables suchas total

amountofDNArecovered.Duetothis,averages,meansorstandarddeviationswerenot

abletobecomputedforresultsincludedinthesestudies,andthereforeameta-analysis

wasnotabletobecarriedout.

ItwasalsodifficulttopinpointastowheretheDNAhadcomefrom,asthepresenceof

DNA on an object or individual could either be a result of secondary DNA transfer or

backgroundDNA.Inthiscase,onlystudieswhocanspecificallytestedforthesecondary

transferofDNAcouldbeincludedinthemeta-analysis,asitwascertainthatthenon-donor

DNAhadcomefromthatspecificevent.However,thisthandecreasedtheexperiment’s

74

applicability to the real world, where now background DNAwas not present, possibly

causinganunrealisticresult.

CONCLUSION

Atthisstage,itcanbeconcludedthatthehypothesis,throughtheexplorationofliterature

andcompletionofameta-analysisthatanappropriateandconciseguidewillbeproduced

todeterminethechancesofasecondarytransfereventoccurringandwhatconditionsare

required,toaidBiologistsinexperttestimony,canberejected.Datapublishedinthe38

studiesincludedforthemeta-analysiswasnotadequateenoughtoperformquantitative

comparisonsbetweenvariables inorder tocombineresults tostrengthenthepowerof

eachstudy.Infuture,itshouldbeconsideredthatarticlespublishedinthefieldofForensic

Sciencedisplaytheirresultsindifferentwaysorgiveaccesstoreadersoftheirrawresults

forfurtherinterpretationpurposes.

75

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META-ANALYSIS OF THE SECONDARY TRANSFER OF DNA · 5 1. INTRODUCTION Deoxyribonucleic acid (DNA) is...

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