Marginal ancestral contributions to atrial fibrillation in the Standardbred racehorse

17 Pages·2017·5.63 MB·English

RESEARCHARTICLE
Marginal ancestral contributions to atrial
fibrillation in the Standardbred racehorse:
Comparison of cases and controls
MeganKraus1,PeterPhysick-Sheard2,LuizF.Brito1,MehdiSargolzaei1,3,Fla´vio
S.Schenkel1*
1 DepartmentofAnimalBiosciences,UniversityofGuelph,Guelph,Ontario,Canada,2 Departmentof
PopulationMedicine,UniversityofGuelph,Guelph,Ontario,Canada,3 TheSemexAlliance,Guelph,
Ontario,Canada
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*[email protected]
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Abstract
AdmissionsofStandardbredracehorses(Std)totheOntarioVeterinaryCollegeTeaching
Hospital(OVCTH)fortreatmentofatrialfibrillation(AF)begantoincreaseintheearly
OPENACCESS 1990s.Thearrhythmiahasbeenshowntohaveamodestheritability(h2’0.15),with
somestallionsappearingassiresorsiresofmaresusedinbreeding(broodmares)of
Citation:KrausM,Physick-SheardP,BritoLF,
SargolzaeiM,SchenkelFS(2018)Marginal affectedhorsesmorefrequentlythanothers.Theobjectiveofthisstudywastodetermine
ancestralcontributionstoatrialfibrillationinthe themarginalgeneticcontributionsofancestorstocohortsofStdaffectedwithAFand
Standardbredracehorse:Comparisonofcasesand
theircontemporarycontrolgroups,andwhethertheseancestorscontributesignificantly
controls.PLoSONE13(5):e0197137.https://doi.
org/10.1371/journal.pone.0197137 moretotheaffectedcohortsthantocontrols.AllStdadmittedtoOVCTHfortreatmentof
AFthatwerebornbetween1993and2007comprisedtheaffectedcasegroup(n=168).
Editor:ClaireWade,UniversityofSydneyFacultyof
VeterinaryScience,AUSTRALIA FiverandomlyselectedracingcontemporariesforeachStdadmitted,assumedtonotsuf-
ferfromthearrhythmia,comprisedthecontrolgroup(n=840).Three-yearoverlapping
Received:September19,2017
cohortswerecreatedforcaseandcontrolhorses,determinedaccordingtoyearofbirth,
Accepted:April26,2018
foratotalof26cohorts.Marginalgeneticcontributionsofancestorstoeachcohortwere
Published:May15,2018 determinedanddifferencesanalyzedforstatisticalsignificanceusingatwo-tailedpaired
Copyright:©2018Krausetal.Thisisanopen t-test,withP(cid:20)0.05consideredsignificant.Themarginalcontributionsof26ancestors
accessarticledistributedunderthetermsofthe weresignificant,with11contributingsignificantlymoretoaffectedcohortsthanthecorre-
CreativeCommonsAttributionLicense,which
spondingcontrols,and15contributingsignificantlymoretocontrolsthanthecorrespond-
permitsunrestricteduse,distribution,and
reproductioninanymedium,providedtheoriginal ingaffectedcohorts.Onestallionandonebroodmarewereveryhighlysignificantto
authorandsourcearecredited. affectedcohortsatP(cid:20)0.001,andninestallionsandthreebroodmareswereveryhighly
DataAvailabilityStatement:Allrelevantdataare significanttocontrolcohortsatP(cid:20)0.001.Therefore,anumberofstallionshavestatisti-
withinthepaperanditsSupportingInformation callysignificantcontributionstothegeneticsofStdaffectedwithAF,whilemanyothers
files.
havestatisticallysignificantcontributionstohealthyStd.Thearrhythmiaappearstobe
Funding:Thisstudywassupportedbythe particularlyprevalentinthedescendantsofonesirefamily.
DepartmentsofPopulationMedicineandAnimal
Biosciences,UniversityofGuelph.Supportwas
alsoprovidedbyStandardbredCanadathrough
theirDataProcessingDivisionandaregratefully
acknowledged.TheSemexAllianceprovided
supportintheformofsalaryfortheco-authorMS.
However,similarlytotheotherfunding
PLOSONE|https://doi.org/10.1371/journal.pone.0197137 May15,2018 1/17
Atrialfibrillationinhorses
organizations,theSemexAlliancedidnothaveany Introduction
additionalroleinthestudydesign,datacollection
Atrialfibrillation(AF)isthemostcommonclinicallysignificantarrhythmiainhorses,andhas
andanalysis,decisiontopublish,orpreparationof
themanuscript.Thespecificrolesoftheauthors amajorimpactonhigh-performanceequineathletes[1,2].Itappearsmostprevalentinrace-
arearticulatedinthe’authorcontributions’section. horses,particularlytheStandardbredracehorse(Std)[3–5].Withthelossintrainingandcom-
petitiontime,potentialforasustaineddecreaseinperformanceability,andtheexpenseand
Competinginterests:Wehavethefollowing
interests.MehdiSargolzaeiisemployedbyThe manneroftreatment,itisreasonabletoassumethatthereisconcernfrombothaneconomic
SemexAlliance.Therearenopatents,productsin andwelfarestandpointwhenconsideringAFinthehorse.
developmentormarketedproductstodeclare.This EvidencepointstowardstherebeingageneticbackgroundtothearrhythmiaintheStd
doesnotalterouradherencetoallthePLOSONE
[6–8].MostinstancesoffamilialAFinhumansaffectyoungerindividualsandhaveanauto-
policiesonsharingdataandmaterials,asdetailed
somaldominantmodeofinheritance.Thisisencouragingforpotentialresearchintothe
onlineintheguideforauthors.
geneticsofAFinthehorse,asitisknownthatinhorsesAFshowsnoclearrelationshipwith
age,soyoungracehorsesaresusceptibletothearrhythmia[9],andmanyoftheknown
geneticdiseasesofthehorsearealsoinheritedinanautosomalrecessiveordominantman-
ner[10].BecauseAFhasbotheconomicandwelfareimplications,effortsshouldbemadeto
decreasethenumberofStdwithapotentialtodevelopthedisease.Twopreviouspublica-
tionsdescribeanalysisofthisdataset[7,8].Inthefirstpaper,breedincidenceofAFwas
described,heritabilityandinbreedingestimatesweredetermined,andthefrequencyof
individualsires(1stgenerationbackinthepedigree)incaseandcontrolgroupswerecom-
pared.ThesecondpaperextendedtheseanalysesandexaminedAF,stratifyingbygaitand
gender.Thesepapersestablishedthebasisforgeneticliabilitytotheconditioninthebreed
andshowedpotentialdifferencesbetweengaitsandgenderswithpossiblylargerheritability
estimatesinmalesandpacers.Thepresentstudyextendstheanalysisfurther,examining
marginalancestralcontributions.
OnewaytoimprovefurtherourunderstandingofthegeneticbasisofAFwouldbetodeter-
minetheancestralgeneticcontributionstothegeneticpoolofthegroupofhorsesthatappear
tobemoresusceptibletothearrhythmiacomparedtothoseofthegroupofcontrol,unaffected
horses.Analysisofapopulationforprobabilitiesofgeneoriginallowsthedeterminationof
ancestralmarginalcontributions[11].Thistypeofanalysiscandiscriminatehorsesorfamilies
moreassociatedwiththeincidenceofAF,allowingforbetterunderstandingofitsgenetic
background.Therefore,theobjectivesofthisstudyweretodeterminethemarginalgenetic
contributionsofancestorstoaffectedandcontrolStandardbredracehorses,andtoidentify
thosehighlycontributingancestorsthathavesignificantlylargercontributionstoaffected
horsesthantocontrols.
Materialandmethods
TheauthorshaveconfirmedthatnoAnimalCareCommitteeapprovalwasnecessaryforthis
particularstudy,asallinformationrequiredwasobtainedfromexistingdatabases.Allclinical
casesweretreatedpriortothisstudy.Owner-informedconsenttotheuseofthedatasetwas
obtainedforuseofthebreedregistrydatasetviathebreederassociation.
Selectionofcaseandcontrolgroups
Thegroupofaffectedanimals(A)andtheircontemporarycontrolgroup(C)isasubsetofthe
groupdescribedpreviouslybyKrausetal.[7].AdmissionsofStandardbredswithAFtothe
OntarioVeterinaryCollegeTeachingHospital(OVCTH)begantomarkedlyincreasestarting
in1993,thusthepedigreesofallcaseandcontrolhorsesbornbetween1993and2007,inclu-
sive,wereexamined.Stratificationsbysexandgaitwerenotusedinordertoavoidproblems
stemmingfromsmallsamplesizes.Ancestralcontributionsweredeterminedforoverlapping
birth-yearcohorts,threeyearsatatime,allowingforthechangeinmarginalcontributionsto
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Atrialfibrillationinhorses
befollowedcloselyovertime.Thefirstcohortsevaluatedwereforthebirthyears1993,1994,
and1995(affected:A93/95,control:C93/95).Subsequentanalyseslookedatbirthyearsof
1994,1995,and1996,(A94/96,C94/96),etc.,uptoandincluding2007(A05/07,C05/07).For
eachcohort,thepedigreesweretracedtotheearliestknownancestors.
Descriptivestatisticsofpedigrees
Inordertoobtainanunderstandingofthecomplexityanddepthofthepedigreesofaffected
andcontrolStd,descriptivedetailswereobtainedthroughCFCsoftware[12],includingaver-
agelevelsofinbreeding,five-generationpedigreecompletenessindex(PCI)[13],numberof
sires,numberofdams,andaveragegeneticrelationshipforStdwithineachcohort.
Marginalgeneticcontributions
Theobjectiveofthisstudywastodetermineifcontributionsofancestorstothecasegroupdif-
feredsignificantlyfromcontributionsofthosesameancestorstothecontrolgroup,therefore
themarginalgeneticcontributionsto13controlcohortsweremeasured,oneforeachaffected
casecohort[11].Ancestorswerechosenbasedontheirgeneticcontributionandare,therefore,
notnecessarilyfoundersofthepopulation.Thehighest-contributingancestorswerechosen
onebyoneinaniterativeprocess,startingwiththehighest-contributingancestor,andpro-
ceedingtothelowest-contributingancestor,withpedigreeinformationupdatedforeachitera-
tion.ThisprocessisexplainedindetailbyBoichardetal.[11].Anancestor’smarginal
contribution,p ,isdefinedasthatcontributionwhichhasnotyetalreadybeenexplainedby
k
then-1previouslyaccounted-forancestors.Whencalculatingthesemarginalcontributions,it
isnecessarytoremovethepossibilityofthecontributionsofanancestorbeingcountedmore
thanonce.Forthefirstmethodofdoingso,then-1selectedancestorsmaybeancestorsofan
individual,k,andthereforethegeneticcontributions,a ofthesen-1individualancestorsare
i
adjustedforaccordingtotheequation:p =q (cid:3)(1-Sa),sothatthemarginalcontributionof
k k i
anancestor,p ,doesnotincludethegeneticcontributionsalreadyexplainedbythen-1ances-
k
tors.Additionally,itispossiblethatsomeofthepreviousn-1ancestorsmaybedescendantsof
individualk,therefore,aftereachmajorancestorhasbeenfound,andtheirrawandmarginal
contributionsdetermined,theinformationontheirsireanddamisdeleted,makingthema
“pseudo-founder,”andavoidingthepossibilityoftheircontributionsbeingattributedto
ancestork.Theprob_origfunctionofthePedigsoftware[14]wasusedtocompletetheseitera-
tivecalculations.
Thenumberofancestorsexplainingacertain%ofthegeneticpool(i.e.,geneticvariability)
wascomputedfromthemarginalcontributionsoftheancestorsgivenbythePedigsoftware
[14].Thisispossible,becausethemarginaldistributionsareorthogonal,i.e.thereisnodouble
countingofcontributions,startingfromthemostcontributingancestorfurtherbackinthe
pedigree.
Inordertoidentifythemostimportantancestorsaccordingtothemagnitudeoftheirmar-
ginalgeneticcontributions,themarginalgeneticcontributionsofancestorstoeachofall13
affectedorcontrolcohortswereaveraged.Ancestorswithanaveragenon-zerocontributionof
0.5%orgreater,thatcontributedtotwoormorenon-overlappingcohorts,wereidentified.
Thecontributionsofthehighest-contributingancestorsidentifiedinthismannerwereana-
lyzedusingapairedtwo-tailedt-test,comparingtheirgeneticcontributionsoverall13three-
yearcohortsofaffectedStdtotheircontributionstothecorrespondingcontrolcohorts.Inall,
41ancestorsweretestedforsignificance.Afalsediscoveryrate(FDR)[15]of5%wasusedto
controlformultiplehypothesestest.
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Results
Thenumberofindividualsineachcohort,aswellasthecompletepedigreesize,includingthe
animalsineachcohort,areshowninTable1.Marginalancestralcontributionsweredeter-
mined,andthemostinfluentialancestorstotheaffectedandcontrolcohortswereexamined
forstatisticalsignificancethroughapairwisetwo-tailedt-testcomparingtheircontributionsto
eachaffectedcohortversusthecorrespondingcontrolcohort.
Descriptivestatisticsofpedigrees
AsseeninTable2,whilenotlarge,therewasanoveralltrendforincreaseininbreedingover
timeinbothaffectedandcontrolgroups,butnoclearpatternexistsforanydifferencesinaver-
ageinbreedingcoefficientbetweenaffectedandcontrolStd.AffectedStdareseentohavea
greaterdegreeofrelatednessthancontrolStd,andcohortsofbothgroupshaveahighdegree
ofpedigreecompletenesswithinfivegenerations.UsingthesameStdaspartofalargerdata-
set,Physick-Sheardetal.[8]reportednoeffectofinbreedingcoefficientontheliabilitytoAF
(P=0.91).Theseauthorsalsoreportedthattherewasasignificantdifferenceinthedistribu-
tionofinbreedingcoefficientsofaffectedandcontrolStd,withcontrolStdfallingmorein
higherinbreedingclassesthanaffectedhorses,indicatingnoeffectofinbreedingonincidence
ofthearrhythmia.
Marginalgeneticcontributions
Marginalcontributionsofindividualancestorstothecontrolcohortsremainedfairlyconsis-
tentovertime(Table3).Thehighestcontributionbyanysingleancestor(ID7),rangedfrom
11.9%toC00/02,to12.7%toC05/07.Thisancestorwasthehighestcontributortoallcontrol
cohortsexceptforcohortC93/95,whenitrankedsecondtoID11.The10highestcontributing
ancestorstothesecondcontrolcohort,C94/96,maintainedtheirhighlevelofcontributionto
allsubsequentcohortswithsomeslightalterationsinorder.Theconsistencyofancestralcon-
tributionstocontrolcohortsisdepictedinFig1A,whichshowsthecontributionsofthefive
highest-contributingancestorstocontrolcohorts.
Marginalcontributionstotheaffectedcohortsvariedmuchmorethanthoseseenwiththe
controlcohorts(Table4).Thehighestmarginalcontributionbyasingleancestor,ID1,was
Table1. Numberofindividualspercohort,andsizeofthecompletepedigreeforeachthree-yearcohortofaffected(A)orcontrol(C)racehorsesusedtodetermine
themarginalancestralcontributionstoeach.
Cohort Sizeofcohort Sizeofpedigree Cohort Sizeofcohort Sizeofpedigree
A93/95 32 1,783 C93/95 160 3,887
A94/96 37 1,873 C94/96 185 4,072
A95/97 41 2,011 C95/97 205 4,389
A96/98 48 2,081 C96/98 240 4,912
A97/99 38 1,881 C97/99 190 4,403
A98/00 32 1,707 C98/00 160 4,210
A99/01 24 1,561 C99/01 120 3,560
A00/02 35 1,836 C00/02 175 4,285
A01/03 46 2,046 C01/03 230 4,821
A02/04 45 1,980 C02/04 225 4,861
A03/05 39 1,830 C03/05 195 4,463
A04/06 27 1,695 C04/06 135 4,019
A05/07 19 1,633 C05/07 95 3,444
https://doi.org/10.1371/journal.pone.0197137.t001
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Table2. Descriptivepedigreestatisticsforeachaffectedandcontrolcohortshowingaverage,maximum,andminimuminbreeding,numberofsires,numberof
dams,averagegeneticrelationship,andaverage5-generationpedigreecompletenessindex(PCI).
Cohort Averageinbreeding Maximuminbreeding Minimuminbreeding Numberof Numberof Average Average
coefficient coefficient coefficient sires dams relationship PCI
A93/95 0.0824 0.1153 0.0394 23 32 0.1841 0.9994
C93/95 0.0820 0.1882 0.0300 129 159 0.1466 0.9963
A94/96 0.0834 0.1302 0.0394 23 37 0.1869 0.9992
C94/96 0.0849 0.1882 0.0300 140 183 0.1463 0.9962
A95/97 0.0837 0.1302 0.0394 28 41 0.1784 0.9988
C95/97 0.0848 0.1882 0.0310 154 204 0.1452 0.9967
A96/98 0.0875 0.1421 0.0550 28 47 0.1811 0.9985
C96/98 0.0857 0.1646 0.0310 178 238 0.1439 0.9971
A97/99 0.0851 0.1421 0.0496 23 38 0.1870 0.9985
C97/99 0.0848 0.1642 0.0368 149 187 0.1445 0.9978
A98/00 0.0875 0.1421 0.0496 18 32 0.2027 0.9988
C98/00 0.0867 0.1645 0.0368 130 158 0.1455 0.9982
A99/01 0.0838 0.1411 0.0440 16 24 0.2251 0.9997
C99/01 0.0896 0.1645 0.0448 105 118 0.1518 0.9986
A00/02 0.0908 0.1501 0.0440 23 35 0.2212 1.0000
C00/02 0.0907 0.1712 0.0325 132 175 0.1522 0.9978
A01/03 0.0898 0.1501 0.0440 23 46 0.2276 0.9997
C01/03 0.0930 0.1831 0.0325 161 230 0.1530 0.9984
A02/04 0.0907 0.1501 0.0610 18 45 0.2473 0.9997
C02/04 0.0947 0.1831 0.0325 159 223 0.1530 0.9984
A03/05 0.0917 0.1488 0.0610 15 39 0.2529 0.9997
C03/05 0.0965 0.1831 0.0375 145 194 0.1550 0.9996
A04/06 0.0994 0.1598 0.0680 15 27 0.2485 1.0000
C04/06 0.0990 0.1639 0.0505 112 134 0.1565 0.9996
A05/07 0.1022 0.1598 0.0680 13 19 0.2547 1.0000
C05/07 0.0987 0.1639 0.0505 82 94 0.1594 0.9997
https://doi.org/10.1371/journal.pone.0197137.t002
20.5%toA03/05.Thisancestorwasinitiallyhighlightedforinterestashehasmanyoffspring
inthegroupofaffectedhorses,andwasthehighest-rankedmarginalcontributorincohorts
A01/03toA05/07.ID1enteredstudin2001,thereforehisfirstcropoffoalswerebornin2002.
ThisisreflectedinthefirstappearanceofID1asacontributingancestortoA00/02.Thevaria-
tioninlevelsofcontributionsofancestorstoaffectedcohortscanbeseeninFig1Bbythecon-
tributionsofthefivehighest-contributingancestorsthatweresignificanttoaffectedcohorts.
Ancestorswereevaluatedfortheirdifferentialmarginalcontributionstotheaffectedand
controlcohorts.Forbothaffectedandcontrolgroups,ancestorswithanaveragecontribution
of0.5%orgreaterineithergroup,andthatcontributedtotwoormorenon-overlapping
cohorts,wereanalyzed.Thisresultedintheidentificationofatotalof41individualancestors
forcomparison.Thedifferenceintheirmarginalgeneticcontributionstoaffectedandcontrol
cohortsweretestedusingatwo-tailedpairedt-test.Theancestors’marginalgeneticcontribu-
tionsareshowninTables3and4andthepairedt-testresultsarepresentedinTable5.
Thepairedt-testrevealed26significantancestors,11significanttoaffectedcohorts,and15
significanttocontrolcohorts.Ofthelatter,12werestallionsandthreewerebroodmares.Of
the11ancestorssignificanttoaffectedcohorts,ninewerestallions,andtwowerebroodmares.
Ofthese26ancestors,23remainedsignificantatanexperimental-wise5%FDR,whilethe
remaining3ancestorsweresignificantata10%FDR.S1andS2Tablesprovideadditional
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Atrialfibrillationinhorses
Table3. Marginalgeneticcontributionstothree-yearcohortsofcontrolhorsesfrom41highest-contributingancestors(Anc.)andoverallaveragecontribution
(Ave.).
Anc. 93/95 94/96 95/97 96/98 97/99 98/00 99/01 00/02 01/03 02/04 03/05 04/06 05/07 Ave.(cid:3)
ID1 - - - - - - - - - - - - - -
ID11 0.125 0.036 0.035 0.032 0.032 0.032 0.032 0.034 0.031 0.031 0.026 0.029 0.023 0.038
ID14 0.086 - - - - - - - - - - - - 0.086
ID4 - - - - - - - - - - - - - -
ID21 0.035 0.108 0.110 0.104 0.108 0.105 0.111 0.111 0.107 0.104 0.100 0.097 0.093 0.099
ID6 0.075 0.087 0.088 0.087 0.087 0.085 0.085 0.085 0.086 0.085 0.086 0.087 0.088 0.085
ID7 0.104 0.121 0.121 0.122 0.120 0.121 0.120 0.119 0.121 0.122 0.125 0.126 0.127 0.121
ID8 0.014 0.013 0.011 0.011 0.011 0.025 0.030 0.043 0.043 0.042 0.041 0.041 0.045 0.028
ID9 0.079 0.094 0.094 0.095 0.093 0.094 0.095 0.093 0.094 0.094 0.097 0.099 0.101 0.094
ID10 0.062 0.075 0.075 0.074 0.074 0.073 0.071 0.073 0.070 0.069 0.068 0.065 0.064 0.070
ID2 - - - - - 0.003 - - - - - - - 0.003
ID12 0.046 0.053 0.053 0.053 0.052 0.052 0.052 0.052 0.053 0.053 0.053 0.053 0.053 0.052
ID13 0.001 0.034 0.034 0.033 0.035 0.033 0.035 0.031 0.030 0.030 0.029 0.027 0.026 0.029
ID3 - - - - - - - - - - - - - -
ID15 - - 0.013 0.021 0.027 0.024 - - 0.017 0.020 0.018 0.019 - 0.020
ID16 0.008 0.004 0.006 0.006 0.008 0.003 0.012 0.012 0.014 0.014 0.015 0.013 0.012 0.010
ID17 0.026 0.028 0.027 0.028 0.026 0.028 0.026 0.025 0.026 0.026 0.030 0.025 0.028 0.027
ID18 0.028 0.030 0.030 0.031 0.029 0.013 0.013 0.013 0.013 0.013 0.013 0.012 0.028 0.020
ID19 0.017 0.019 0.018 0.019 0.018 0.029 0.029 0.028 0.030 0.031 0.033 0.026 0.014 0.024
ID20 0.022 0.020 0.020 0.019 0.019 0.016 0.019 0.019 0.020 0.019 0.020 0.019 0.017 0.019
ID5 - - - - - - - - - - - - - -
ID22 - - - - - 0.009 - - - - - - - 0.009
ID23 0.010 0.012 0.014 0.016 0.014 0.012 0.012 0.012 0.014 0.014 0.015 0.016 0.020 0.014
ID24 - - - - - - 0.006 - - - - - - 0.006
ID25 - - - - - - - - - - - - - -
ID26 - - - - - - - - - - - - - -
ID27 0.007 0.007 0.007 0.007 0.007 0.007 0.007 0.006 0.006 0.006 0.006 - 0.006 0.007
ID28 0.013 0.016 0.017 0.019 0.018 0.020 0.018 0.018 0.021 0.023 0.025 0.025 0.023 0.020
ID29 0.018 0.017 0.017 0.014 0.014 0.013 0.011 0.011 0.011 0.011 0.011 0.010 0.013 0.013
ID30 0.013 0.015 0.010 0.012 0.011 0.019 0.020 0.019 0.020 0.021 0.023 0.034 0.036 0.019
ID31 0.006 0.007 0.007 0.007 0.006 0.007 0.007 0.006 0.006 0.005 0.005 0.004 0.006
ID32 - - - - - - - - - - - - - -
ID33 - - - - - - - - - - - - - -
ID34 - - - - - - - - - - - - - -
ID35 - - - - - - - - - - - - - -
ID36 0.002 0.002 0.002 0.002 0.002 0.001 0.001 0.001 0.001 0.001 0.001 0.002 0.001 0.001
ID38 0.011 0.010 0.003 0.004 0.005 0.004 0.020 0.018 0.005 0.004 - 0.006 - 0.008
ID41 0.007 0.008 0.008 0.008 0.008 0.007 0.007 0.007 0.008 0.008 0.008 0.009 0.009 0.008
ID37 0.007 0.008 0.008 0.008 0.008 0.007 0.007 0.007 0.007 0.007 0.008 0.008 0.008 0.007
ID40 0.007 0.008 0.008 0.008 0.008 0.007 0.007 0.007 0.007 0.007 0.007 0.007 0.007 0.007
ID39 0.004 0.005 0.005 0.006 0.005 0.005 0.005 0.005 0.005 0.004 0.005 0.006 0.006 0.005
(cid:3)Ancestorswithnovaluesinallcolumnsareancestorsthathadmarginalcontributionstotheaffectedgroup,butnottothecontrolgroup.
https://doi.org/10.1371/journal.pone.0197137.t003
informationonsignificantstallionsandbroodmares(26ancestors),includingyearofbirth
andgait.Statisticalresultsforall41ancestorstestedareshowninTable5.
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Fig1.Marginalgeneticcontributionsofthefivehighest-contributingancestorssignificanttocontrol(Fig1A)andaffected(Fig1B)cohorts,selectedaccordingto
largestaveragenon-zerocontribution.
https://doi.org/10.1371/journal.pone.0197137.g001
Foreachcontrolcohort,thenumberofancestorsnecessarytoexplain50%ofthegenepool
was6.Thesamewastruefor5oftheaffectedcohorts.Theremaining8,A93/95,A99/01,A00/
02,A01/03,A02/04,A03/05,A04/06,andA05/07,neededonly5ancestorstoexplain50%of
thegenepool.Thefewestnumberofancestorstoexplain75%ofthegenepoolforacontrol
cohortwas14,fromC03/05toC05/07.Only10ancestorswerenecessarytoreach75%ofthe
genepoolincohortsA02/04andA03/05.Duetothepresenceofunknownancestorsinsome
ofthepedigrees(theStdstudbookwasnotclosedtooutsidehorsesuntil1973),itmaynotbe
possibletoexplain100%ofgeneticpool.Therefore,thenumberofancestorsnecessaryto
explain99%ofgeneticvariabilitywasobtained.Asfewas43ancestorsaccountedfor99%of
geneticvariabilityintheA05/07group,while129wastheminimumrequiredforC05/07.This
differencemaybeduetothesmallernumberofindividualsintheaffectedcohorts.Eighty-two
ancestorsexplainalmosttheentirevariabilityforA96/98,and199forC96/98,whicharethe
largestcohorts.S3Tablepresentscompletedetailsregardingthenumberofancestorsexplain-
ing50%,75%,and99%ofthegeneticsofeachofthe26affectedandcontrolcohorts,andthe
greatestmarginalancestralcontributiontoeach.Apictorialrepresentationoftheseresultsis
showninFig2.
Afterdeterminingtheancestorsthatweresignificanttoaffectedcohorts,thefrequencyof
theirappearanceinthefive-generationpedigreesofaffectedStdwasexamined.Anumberof
affectedStdhadstallionsID4,ID11,orID15appearthroughboththeirsireanddam.These
threeparticularancestorsappearatleastoncein84.3%,70.6%,and10.8%ofallcasesofAFat
OVCTH,respectively.FurtherdataonID4,ID11,andID26areshowninFig3.Oftheremain-
ingancestorssignificanttoaffectedcohorts,noneappearthroughthesireordamofaffected
horsesmorethanonce.However,ID8appearsinthefive-generationpedigreeof44.6%ofdis-
easecases,whereasID1appearsonlyasasireofcasehorses.Fig4showsdatafortheremaining
8significantancestors,andthenumberoftimeseachappearsthroughthesireordamofacase
horse.Interestingly,ID4andID26appearmorefrequentlythroughthedamofaffected
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Table4. Marginalgeneticcontributionstothree-yearcohortsofaffectedhorsesfrom41highest-contributingancestorsandoverallaveragecontribution.
Anc. 93/95 94/96 95/97 96/98 97/99 98/00 99/01 00/02 01/03 02/04 03/05 04/06 05/07 Ave.
ID1 - - - - - - - 0.010 0.174 0.200 0.205 0.148 0.158 0.149
ID11 0.038 0.128 0.142 0.147 0.144 0.141 0.099 0.101 0.028 0.030 0.026 0.119 0.110 0.096
ID14 - 0.094 - 0.095 0.100 - - - - - - 0.066 0.071 0.085
ID4 - - 0.083 - - - - 0.084 - - - - - 0.083
ID21 0.127 0.047 0.086 0.038 0.046 0.094 0.141 0.142 0.115 0.093 0.088 0.033 0.035 0.083
ID6 0.116 0.102 0.057 0.099 0.090 0.085 0.051 0.039 0.066 0.075 0.074 0.067 0.075 0.077
ID7 0.084 0.069 0.100 0.066 0.064 0.061 0.080 0.070 0.089 0.055 0.054 0.043 0.051 0.068
ID8 0.032 0.042 0.047 0.036 0.041 0.106 0.129 0.122 0.045 0.043 0.044 0.093 0.092 0.067
ID9 0.087 0.076 0.068 0.070 0.069 0.059 0.051 0.049 0.067 0.055 0.053 0.048 0.057 0.062
ID10 0.088 0.076 0.019 0.074 0.072 0.067 0.051 0.017 0.074 0.062 0.058 0.045 0.049 0.058
ID2 0.019 0.021 - - - - 0.022 0.024 0.044 0.117 0.128 0.081 0.006 0.051
ID12 0.050 0.036 0.041 0.042 0.035 0.033 0.028 0.027 0.033 0.021 0.030 0.024 0.028 0.033
ID13 0.038 - 0.031 0.000 - 0.042 0.036 0.041 0.027 0.032 0.023 - - 0.030
ID3 - - 0.016 0.011 0.016 0.025 0.069 - - - - - - 0.027
ID15 0.026 0.007 0.017 0.019 0.026 0.020 - 0.017 0.023 0.027 0.026 0.029 0.031 0.022
ID16 0.019 0.023 0.005 - - - 0.004 0.056 0.018 0.012 0.014 0.019 0.024 0.019
ID17 0.023 0.024 0.024 0.019 0.022 0.015 0.012 0.016 0.014 0.013 0.013 0.016 0.020 0.018
ID18 0.024 0.025 0.010 0.021 0.020 0.015 0.012 0.015 0.016 0.015 0.015 0.017 0.019 0.017
ID19 - - 0.025 - - - - 0.008 - - - - 0.012 0.015
ID20 0.013 0.012 0.017 0.017 0.022 0.016 0.013 0.017 0.011 0.011 0.010 0.014 0.016 0.015
ID5 0.002 - 0.002 0.026 0.012 0.026 - - - - - - - 0.013
ID22 - - - 0.001 - 0.012 0.025 - - - - - - 0.012
ID23 - 0.009 0.013 0.010 0.015 - - - - - - - - 0.012
ID24 0.005 - - - 0.016 0.019 - 0.006 0.007 - - - - 0.011
ID25 - 0.007 0.006 - - - 0.017 - - - - - - 0.010
ID26 0.017 - - - - - - - 0.007 0.006 - - 0.009 0.009
ID27 0.012 0.012 0.003 0.012 0.009 0.006 - - 0.009 0.008 0.008 0.010 - 0.009
ID28 0.011 0.010 0.012 0.008 0.007 - - 0.007 0.007 0.005 - - 0.010 0.008
ID29 0.010 0.011 0.008 0.007 0.009 0.006 - 0.007 0.008 0.008 0.008 - - 0.008
ID30 0.009 0.016 0.008 0.007 0.007 0.009 - 0.010 0.004 - 0.002 - - 0.008
ID31 - 0.001 0.009 0.013 0.004 0.004 0.005 - - 0.006 0.006 0.009 0.009 0.007
ID32 0.006 - - - 0.005 0.007 0.007 - - - - - - 0.006
ID33 0.003 0.003 0.011 0.009 0.002 - - - - - - - - 0.006
ID34 0.008 0.009 0.001 - - - - 0.008 - - 0.002 - - 0.006
ID35 0.002 0.002 - 0.007 0.010 0.010 0.009 0.005 0.003 0.002 - - - 0.005
ID36 0.007 0.006 - 0.006 0.005 - - - 0.005 0.005 0.003 - - 0.005
ID37 0.005 - 0.005 0.005 0.004 - - 0.004 0.005 0.005 - - - 0.005
ID38 - - 0.001 0.002 0.002 0.002 0.013 - - - - - - 0.004
ID39 - - 0.003 - - - - - - - - - - 0.003
ID40 0.006 - 0.004 0.003 - - - - 0.002 0.000 - - - 0.003
ID41 - - 0.005 - - - - - - 0.000 - - - 0.003
https://doi.org/10.1371/journal.pone.0197137.t004
racehorsesthanthroughtheirsire.Percentageoffive-generationpedigreesofcasehorsesin
whichasignificantancestorispresentisshowninFig5.Lastly,afterobservingtheregularity
withwhichindividualsfromaparticularsireline(ID1,ID8,ID42)wereappearinginthecase
group,apedigreediagramforthisfamily,Fig6,wasconstructedusingPedigreeViewer[16].
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Table5. Pairwisetwo-tailedt-testsforthe41ancestorswiththehighestaveragemarginalcontributionstotwoormorenon-overlappingcohorts.
Ancestor DF t P FDR Group
ID1 5 5.0863 <0.01 (cid:3) Affected
ID11 12 3.3826 <0.01 (cid:3) Affected
ID4 1 151.7273 <0.01 (cid:3) Affected
ID8 12 4.354 <0.001 (cid:3) Affected
ID2 9 3.0727 <0.02 (cid:3) Affected
ID15 11 3.1227 <0.01 (cid:3) Affected
ID26 3 3.7823 <0.05 # Affected
ID32B 3 14.4752 <0.001 (cid:3) Affected
ID33 4 3.0569 <0.05 # Affected
ID34B 4 3.3738 <0.05 # Affected
ID35 8 4.6647 <0.002 (cid:3) Affected
ID7 12 -9.4784 <0.001 (cid:3) Control
ID9 12 -7.3222 <0.001 (cid:3) Control
ID12 12 -7.3755 <0.001 (cid:3) Control
ID17 12 -7.2343 <0.001 (cid:3) Control
ID19 12 -6.4782 <0.001 (cid:3) Control
ID20 12 -4.1063 <0.002 (cid:3) Control
ID23 12 -6.0075 <0.001 (cid:3) Control
ID28 12 -7.024 <0.001 (cid:3) Control
ID29 12 -7.1564 <0.001 (cid:3) Control
ID30 12 -4.2141 <0.002 (cid:3) Control
ID38 10 -4.2976 <0.002 (cid:3) Control
ID41B 12 -17.475 <0.001 (cid:3) Control
ID37 12 -7.0793 <0.001 (cid:3) Control
ID40B 12 -11.2122 <0.001 (cid:3) Control
ID39B 12 -17.6036 <0.001 (cid:3) Control
ID14 5 1.9418 N.S. -
ID21 12 -1.209 N.S. -
ID6 12 -1.372 N.S. -
ID10 12 -1.9819 N.S. -
ID13B 12 -1.3826 N.S. -
ID3 4 2.5778 N.S. -
ID16 12 1.3197 N.S. -
ID18 12 -1.6611 N.S. -
ID5 4 2.4813 N.S. -
ID22 2 1.2627 N.S. -
ID24 4 2.5977 N.S. -
ID25 2 3.0586 N.S. -
ID27B 12 0.5217 N.S. -
ID31B 12 -0.3303 N.S. -
ID36B 12 1.7789 N.S. -
N.S.:contributionsarenotsignificantlydifferenttoaffectedorcontrolcohorts;
B:denotesbroodmares.
FDR:significantatanexperimental-wise5%((cid:3))or10%(#)falsediscoveryrate.
https://doi.org/10.1371/journal.pone.0197137.t005
Inthisstudy,themajorityofthestallionsthatcontributedsignificantlytoaffectedcohorts
arefrompacingbloodlines,butID26isfromtrottingbloodlines.Interestingly,manyofthe
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Fig2.Numberofancestorsexplaining50,75,and99%ofthegenepoolseenin26cohortsofcontrol(C)oraffected(A)racehorses.
https://doi.org/10.1371/journal.pone.0197137.g002
siresfrompacinglineagearerelatedtoeachotherwithinfewerthanfivegenerations.ID1,
ID2,ID33,ID34,andID35alldescendfromID11.Additionally,ID1,ID8,ID33,andID35
descendfromID4.Inter-relationshipsbetweentheseancestorsareshowninFig7.Thesires
whichcontributedmoresignificantlytocontrolsthantoaffectedhorsestendtoappearmore
frequentlyasgreat-grandparents,orevenfurtherbackinthepedigreesofaffectedandcontrol
horses.Theyarerepresentativeofbothtrottingandpacinglineages.ThebroodmareID32has
twoaffectedoffspring,oneofwhichissiredbyID33,himselfsignificanttoaffectedcohorts.
Additionally,broodmareID34issignificant,andisthedamofthestallionID3,notedinKraus
etal.[7]forhislargenumberofaffectedprogeny.
Discussion
Animportantinformationformanaginggeneticdiversityofapopulationistheancestors’
marginalcontributions.Forincreasingormaintainingdiversity,descendantsofhighlycon-
tributingancestorsmightbeusedlessforbreedinganddescendantsfromlesscontributing
ancestorscouldbeusedmoreintensively.ThesamestrategymightbeusedintheStdhorse
population,butaimingtomanageAFincidence.ThePedigprogramdevelopedbyBoichard
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