Pedigree reconstruction of wine and table grape crossbreeds created in Italy by Giovanni Dalmasso

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Contents lists available atScienceDirect

Scientia

Horticulturae

j o u r n a l h o m e p a g e :w w w . e l s e v i e r . c o m / l o c a t e / s c i h o r t i

Pedigree

reconstruction

of

wine

and

table

grape

crossbreeds

created

in

Italy

by

Giovanni

Dalmasso

Stefano

Raimondi

a,∗,1

_,

_Antonio

_Carlomagno

b,1

_,

_Paola

_Ruffa

a

_,

_Sara

_Oglietti

b

_,

Vittorino

Novello

b

_,

_Anna

_Schneider

a

a_National_Research_Council_of_Italy_(CNR),_Institute_for_Sustainable_Plant_Protection_(IPSP)₋_Unit_Grugliasco,_Largo_P._Braccini_2,₁₀₀₉₅_Grugliasco,_TO,_Italy b_Department_of_Agriculture,_Forest_and_Food_Sciences_(DiSAFA),_University_of_Turin,_Largo_P._Braccini_2,₁₀₀₉₅_Grugliasco,_TO,_Italy

a

r

t

i

c

l

e

i

n

f

o

Articlehistory:

Received27October2016

Receivedinrevisedform24January2017 Accepted28February2017 Keywords: Microsatellites VitisviniferaL. Genotyping Parentageanalysis SSR

a

b

s

t

r

a

c

t

Startingin1931,GiovanniDalmassocarriedoutintensegrapevinebreedingactivity,creatingmorethan

100wineandtablegrapecrosses.HesoughttocontributetotheeconomicdevelopmentofItalian

viticulture,butalsoleftconsiderablegeneticmaterialforfurtherbreedingprograms.

Owingtothecurrentstronginterestinthegeneticimprovementofgrapevines,thepedigreesof

vari-etiesreleasedbybreedersmustbedetermined.Theaimofthisstudywastogeneticallycharacterizeand

verifythedisclosedpedigreesofDalmasso’scrosses(IDs).Nuclearmicrosatelliteproﬁles(n-SSR)of42

IDaccessionsand22genotypesdeclaredasparentswereobtainedat22loci.Bycross-validationofallele

size,declaredparentageswereveriﬁed.Whenoneorbothdisclosedparentswerefoundtobe

incor-rectduetoinconsistentgeneticdata,putativeparent(s)weresoughtviaSSRproﬁlecomparisonwithin

thegrapevinemoleculardatabaseofCNR-InstituteforSustainablePlantProtection,andtheprobability

estimatedviaIDENTITYv.4.0software.

Throughmicrosatelliteanalysis,threeduplicatedgenotypeswerediscoveredandtwentyIDparentages

outof39wereconﬁrmed.In13IDs,oneparentwasincorrect,in2IDs,bothparentswereinconsistent

withmicrosatelliteproﬁles,andin4IDsthepedigreecouldnotbeveriﬁedsincethepollendonorwas

notavailable.Apartfrominvalidatedcrosseslikelyduetopollencontamination,5accessionswere

mis-labelled,eitherwhenbothparentswereinvalidatedorwhentwospecimensofthesameoffspringwere

differentlylabelled.Veriﬁcationofbreeder’sdeclaredparentagesrevealed43%ofinvalidatedpedigrees

withintheinvestigatedwineandtablecross-breeds.Theresultsprovideadditionalinsightintograpevine

availablediversity,andmayaidthedevelopmentoffurtherbreedingprograms.

1. Introduction

VegetativepropagationofVitisspp.varietieshasbeenusedfor centuries,preservingthecharactersofthefounderplants.Except perhapsintheearlystagesofdomestication,orincasesofseed transportationbymigratingpopulations,reproductionfromseed hastendedtobeavoided,becauseoftheuncertaintraitsexhibited by descendants. The grapevine is a highly heterozygous cross-pollinatingspecies(Thisetal.,2006).However,cross-breedingis veryadvantageousfordevelopingvarietieswithfavourabletraits, especiallywhenpollinationcontrolandoffspring’sstrictselection areapplied.Asfarasis known,theﬁrstsuccessesingrapevine

∗ Correspondingauthor.

E-mailaddress:[email protected](S.Raimondi).

1 _These_authors_contributed_equally_to_the_work_and_share_ﬁrst_authorship.

genetic improvement by deliberate crossing date to the early nineteenthcentury,whenLouisBouschetobtainedthetwo ﬂesh-colouredvarieties‘PetitBouschet’and‘GrosBouschet’(laterused todevelopotherﬂesh-colouredwinegrapevarieties)(Alabouvette, 1936),andwhenotherbreeders,suchasFosterintheUKandVibert inFrance,releasedimprovedtablegrapecultivars.

After these pioneering successes, intense breeding activity, startingfromthemid-nineteenthcentury,waschieﬂydirectedat obtaininghybrids(mainlyfromNorthAmericanspeciesandVitis viniferaL.)resistanttofungaldiseasesandphylloxera.Apartfrom breeding for resistance, during thelast 150 years many Italian geneticists(Bruni,Cosmo,Dalmasso,Manzoni,Pirovano,Prosperi, Rigotti,Terzi, andothers)aswellasthosefromothercountries (Branas,Gargiulo,Mathiasz,Müller,Olmo,Thomson,Truel,Vidal, toname a few)concentrated theircross-breedingprograms on VitisviniferaL.,aimingtoobtainnewtablegrapeswith

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126 S.Raimondietal./ScientiaHorticulturae219(2017)125–130 ingfeatures,andnewwinegrapecultivarsfromwhichtoproduce

appealingmarketablewines.

Amongthe scientists mentioned, GiovanniDalmasso started a cross-breedingprogram onwine and tablegrapesin 1931at theConeglianoViticultureExperimentalStation(Veneto, North-EasternItaly).TheprogramwascontinuedattheUniversityofTurin (Piedmont,North-WesternItaly). Theprogram concerningtable grapesaimed:a)toobtainimprovedgenotypesfromtraditional varieties;b)tomergethegrapequalitycharacteristicsfromtwoor moregenotypes(berryshape,colorandﬂavour)withresilienceand adaptability,c)tocreatenewgenotypessuitedalsotothecooler climateof NorthernItaly. Thewine grapeprogram was chieﬂy concentratedondevelopingnewcrossbreedscombiningpositive technologicalfeatures.AccordingtoProf.Dalmasso’soriginalnotes, duringtwentyyearsofactivitymorethan100wineandtablegrape crosseswereselected,leavingalargebodyofgrapediversityand considerablegeneticmaterialforfurtherbreeding.

Amongthesenewcultivars,14winegrapevarietiesand6table grapegenotypes are includedin theItalian NationalGrapevine VarietyCatalogue.Someofthem,likethewinegrape‘Albarossa’ producingremarkablewines,currentlyenjoythefavourof grow-ersandaregreatlyappreciatedbythemarket.Othershavenever emergedfromcollectionsorexperimentaltrials.

Amongthemostcommonlyusedmolecularmarkersforgenetic analyses in grapevines, microsatellite alleles are inherited in Mendelianco-dominantsegregation,confirmingtheirsuitability forgenomemappingandtoinvestigatehereditabilityand culti-varparentage (Thomas etal.,1994; Sefcetal., 2009).Theycan thusbeusedtoconfirmorinvalidatebreeder’sinformationonthe pedigreeofabreed.Beforetheuseofmolecularmarkers,the puta-tiveparentageofnewgrapecultivarswasretrievedfrombreeders’ notes,whichcouldbeincompleteor inaccurate.Severalstudies have usedmicrosatellite markers to clarifythe parentage rela-tionshipsbetweengrape cultivars, eithertraditional or modern (Thomasetal.,1994;Ciprianietal.,2010;Lacombeetal.,2013).In grape-breedingprograms,thefemaleparentiscertain,whilethe putativepollendonormightbemistakenbecauseof contamina-tionduringpollination.Thisopensthepossibilityofamaleparent notcorrespondingtothereportedpedigree(Dettweileretal.,2000; Ibà ñezetal.,2009;Vargasetal.,2009;Lacombeetal.,2013).Insome cases,duetomistakenvarietalidentity,thefemaleparentmayalso beincorrect(Akkaketal.,2007).In connectionwithDalmasso’s crossesinvolving‘Nebbiolo’asparent,TorelloMarinonietal.(2009)

discoveredthat,insteadofthetrue‘Nebbiolo’,Dalmasso acciden-tallyused‘NebbiolodiDronero’,asynonymfor‘Chatus’thatdiffers considerablyfrom‘Nebbiolo’.

Becauseof the aboveerrors, this study aimedto clarifythe pedigreesofbreedsobtainedbyProf.Dalmassothatarestill main-tainedincollectionsorcommerciallyexploited.Thesecondaryaim wastogeneticallycharacterizethesematerials,offeringreference geneticﬁngerprintsfor theuseofbreeders,grape growers,and thewineindustry.Duetotheeconomicrelevanceoftablegrape breeding,thereisworldwideinterestinknowingvarieties’correct pedigrees,becausethis informationhelpsbreederstoplantheir cross-breedingprograms.

Astothegrapevarietiesdiscussed,manyofthem,especially thetablegrapes,aresuitableforuseinfurtherbreedingprograms, becausetheyexhibitimprovedtraitsincomparisontotheir par-ents(Carlomagnoetal.,2014).Atthesametime,newvarietiesare morelikelytobeacceptedanddevelopedinthemoredynamictable grapemarket.Inrecentyears,muchresearchactivityisongoing attheDepartmentofAgriculture,ForestandFoodSciences (Uni-versityofTurin,Italy),aimedtocharacterizetheagronomicand qualitativebehaviourofDalmasso’stablegrapecrossesinvarious environments,intheattempttoencouragetheiruseatthelocal andnationallevel.

Apartfromtheirgeneticvalueasasourceofdiversity,newwine grapecultivarscouldplay aroleinthemoretraditionalistwine market,throughfavourablefeaturessuchasadaptabilitytoclimate change,orbyofferinganoriginal,appealingﬂavour.

2. Materialsandmethods 2.1. Plantmaterial

TheSSRproﬁlesof42Dalmassointerbreeds(IDs)andof22 cul-tivarsdeclaredasparentswereobtained.SamplesfrombothIDs anddeclaredparentsweretakenfromﬁeldcollections.Inonecase (‘Pirovano62’)itwasimpossibletoretrievesamplesofthedesired accession,becauseitwaslackinginknowncollections.

The varieties (IDs) under investigation were described in terms of vine morphological features during the AGERProject, 2010–2104, “An Italian Vitis database with multidisciplinary approach,forexploitationandvalorisationoftheregional geno-types”, employing themajor OIV descriptorsselected fromthe EuropeanVitisDatabase(http://www.eu-vitis.de),andcompared withpublishedreferences.

Mostoftheplantmaterialwastakenfromtwocollections main-tainedbytheUniversityofTurin,locatedatChieri(TurinProvince; plantedin1975)andatAlba(CuneoProvince;plantedin1984); 22accessionswerefromtheCNR-IPSPcollectionlocatedat Grin-zaneCavour(CuneoProvince;plantedin1992);twosampleswere kindlyprovidedbyCREA-VIT(Conegliano,Italy).Aftercheckingthe uniformityofallthevinesbymorphology,foreachaccessionone vinewaschosen,fromwhichtotakea sampleofyoungleaves, andthevinewaslabelledforfurthercontrols.Samplesconsisting of3–4youngleaveswerethenstoredat−80◦_C_until_extraction;

DNAextractionwasdonebytheThomasandScottprotocol(1993), slightly modiﬁed. Table 1 lists the 42 IDs analysed with their declaredparents.

2.2. SSRanalyses

ExtractedDNA sampleswereamplifiedbyPolymeraseChain Reaction(PCR)usingprimerslabelledwithfourdifferent fluores-centdyes(Fam,Ned,Pet,Hex).ThePCRreactionswererunina BIO-RADT100thermalcycler,withthefollowingthermalprofile: onecycleat95◦Cfor3min,followedby28cyclesat95◦Cfor30s each,52◦Cfor45s,72◦Cfor90s,andafinalstepof30minat72◦C. Amplificationproductswereanalysedona3130Genetic Ana-lyzer capillary sequencer (Applied Biosystems, Foster City, CA, USA).TheinternalGeneScansizestandard500LIZ(Applied Biosys-tems)wasincludedineachrun.Allele sizesintheoutputwere determinedusing GeneMapperv.4.0 software(Applied Biosys-tems),andallelesweredesignatedbytheirsizeinbasepairs(bp). Withregardtothemarkersused,atthefirststep,allIDsunder studyandtheirputativeparentswereanalysedbythe9SSR mark-ersselectedbytheinternationalscientificcommunityforcommon use(Thisetal.,2004;Grapegen06:http://www1.montpellier.inra.

fr/grapegen06/technical/index.html): VVS2 (Thomas and Scott,

1993),VvMD5,VvMD7(Bowersetal.,1996),VvMD25,VvMD27, VvMD28,VvMD32(Bowersetal.,1999),VrZag62andVrZag79(Sefc etal.,1999).

Declaredparentageswereverifiedbycross-validationofallele size.Whenoneorbothdisclosedparentswerefoundtobe incor-rect,duetoinconsistentgeneticdata,theputativeparent(s)were soughtviaSSRprofilecomparisonwithinthegrapevinemolecular databaseofCNR-InstituteforSustainablePlantProtection (CNR-IPSP),whichcontains850uniquegenotypesofEuropeancultivars (unpublished).Atthesecondstage,allinvestigatedIDs,their par-entsvalidatedbythefirst9markersused,andtheputativeparents

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Table1

The42investigatedcultivarsissuedfromDalmasso’scrosses.

Cultivarcode Cultivarname Utilization Berrycolora _Parents_according_to_breeder_notations_(variety_names_according_to_VIVCb₎

II-26 Vega Wine B Furmint×MalvasiaCosolo(Malvasiaistriana)

II-32 Fubiano Wine B Furmint×Trebbianotoscano

III-3 Table B Moscatod’Amburgo(MuscatHamburg)×Reginabianca(AfusAli) III-32 Table B Moscatod’Amburgo(MuscatHamburg)×Reginabianca(AfusAli) III-34 Franca Table N Moscatod’Amburgo(MuscatHamburg)×Reginabianca(AfusAli)

IV-6 Table N Chasselasrosa(Chasselasrose)×PerladiCsaba(CsabaGyoengye)

IV-28 Cornarea Wine N Barbera×Nebbiolo

IV-31 Soperga Wine N Nebbiolo×Barbera

V-1 Table N Chasselasrosa(Chasselasrose)×PerladiCsaba(CsabaGyoengye)

VI-3 Emilia Table B Bicane×Reginabianca(Afusali)

VI-6 Table B Bicane×Reginabianca(Afusali)

VI-24 Table B Bicane×MoscatodiTerracina

VII-21 S.Martino Wine N Nebbiolo×Dolcetto

VIII-1 Table B Bicane×Moscatod’Amburgo(MuscatHamburg)

VIII-5 Bionda Table B Bicane×Moscatod’Amburgo(MuscatHamburg)

VIII-10 Table N Bicane×Moscatod’Amburgo(MuscatHamburg)

IX-2 Table B Schiavone(Schiavagrossa)×Zibibbo(MuscatofAlexandria)

X-4 Wine B Verdiso×Maddalenareale(Madeleineroyale)

X-10 Wine B Verdiso×Maddalenareale(Madeleineroyale)

X-12 Sirio Wine B Verdiso×Maddalenareale(Madeleineroyale)

XI-2 Table B Moscatod’Amburgo(MuscatHamburg)×Reginabianca(AfusAli)

XI-5 Table Rs Moscatod’Amburgo(MuscatHamburg)×Reginabianca(AfusAli)

XII-26 Wine B Furmint×Malvasiatrevigiana(Malvasiabiancalunga)

XII-30 Wine B Furmint×Trebbianotoscano

XII-37 Bussanello Wine B Rieslingitalico(Welschriesling)×Furmint

XII-40 Wine B Rieslingitalico(Welschriesling)×Rieslingrenano(Rieslingweiss) XIII-11 Covè Wine B Hàrslevelü×Malvasiatrevigiana(Malvasiabiancalunga)

XIV-4 Wine N Grignolino×Sangiovesegrosso(Sangiovese)

XIV-15 Wine N Grignolino×Sangiovesegrosso(Sangiovese)

XV-29 Nebbiera Wine N Nebbiolo×Barbera

XV-31 Albarossa Wine N Nebbiolo×Barbera

XV-34 S.Michele Wine N Nebbiolo×Barbera

XVI-8 Valentino Wine Rg Nebbiolo×Dolcetto

XVII-25 Passau Wine N Dolcetto×Nebbiolo

XVIII-3 Viola Table N Moscatod’Amburgo(MuscatHamburg)×I.P.62(Pirovano62) XVIII-12 Liana Table N Moscatod’Amburgo(MuscatHamburg)×I.P.62(Pirovano62) XVIII-21 Giovanna Table N Moscatod’Amburgo(MuscatHamburg)×I.P.62(Pirovano62) XVIII-24 Teresita Table B Moscatod’Amburgo(MuscatHamburg)×I.P.62(Pirovano62)

a_B₌_white,_Rg₌_red,_N₌_black,_Rs₌_rose.

b_Vitis_{International}_Variety_Catalogue:_{http://www.vivc.de/index.php.}

retrievedfromtheCNR-IPSPinternalSSRdatabase,wereanalysed byafurther13microsatelliteloci.Theseloci (belongingto var-iouslinkagegroups,differentfromthose oftheprevious9SSR) were:VvMD21,VvMD24,VvMD26(Bowersetal.,1999);VrZag64 (Sefcetal.,1999);VVIb01,VVIn73,VVIp31,VVIq52(Merdinoglu etal.,2005);VMC2h4,VMC7f2,VMC5g8,VMC1e8,andVMC3d12 (VitisMicrosatelliteConsortiummanagedbyAgrogeneSA,Moissy Cramayel,France).All19grapevinechromosomeswerecovered bythe22utilisedmarkers.Parentageconsistencywasevaluated statisticallyon290geneticproﬁlesobtainedat22SSRlocifrom: a)theinvestigatedcross-breeds(42accessions),b)thedeclared andputativeparents(21),andc)afurther227V.viniferacultivars. Parentageprobabilityandthemaingeneticparameterswere esti-matedviaIDENTITYv.4.0software(developedbyWagnerandSefc, 1999).IDENTITYcomputesthelikelihoodratios(bothcumulative andwithanupperconﬁdencelimitof95%)ontheallelefrequencies ofaproposedparentageversusotherpossibilities.PIC (Polymor-phismInformationContent)wascalculatedviaPOWERMARKER version3.25software(http://www.powermarker.net).

3. Resultsanddiscussion

TheuniqueSSRproﬁlesoftheinvestigatedIDsandtheir par-entsarereportedintheSupplementaryMaterial.Table2givesthe

Table2

Geneticparametersofa290-samplesetgenotypedwith22SSRloci.

LOCUS Na He Ho PI PIC VvMD5 9 0.822 0.855 0.054 0.800 VvMD7 13 0.798 0.821 0.066 0.773 VvMD25 11 0.766 0.772 0.093 0.727 VvMD27 8 0.799 0.769 0.070 0.770 VvMD28 15 0.876 0.907 0.028 0.864 VvMD32 13 0.797 0.841 0.065 0.774 VvS2 12 0.836 0.855 0.045 0.819 VrZAG62 10 0.845 0.890 0.043 0.826 VrZAG79 11 0.837 0.865 0.043 0.821 VvMD21 5 0.549 0.572 0.250 0.502 VvMD24 6 0.684 0.724 0.156 0.627 VvMD26 8 0.673 0.755 0.160 0.619 VrZAG64 15 0.839 0.879 0.045 0.820 VMC1e8 12 0.839 0.827 0.046 0.819 VMC2h4 16 0.787 0.759 0.068 0.764 VMC3d12 10 0.699 0.710 0.126 0.664 VMC5g8 8 0.710 0.762 0.127 0.667 VMC7f2 6 0.600 0.654 0.195 0.565 VVIb01 5 0.686 0.731 0.156 0.628 VVIn73 6 0.289 0.259 0.517 0.277 VVIp31 12 0.884 0.921 0.025 0.873 VVIq52 7 0.673 0.717 0.168 0.611 Cumulative 3.23×10−24 0.710 Na=numberofalleles,He=expectedheterozygosity,Ho=observedheterozygosity,

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128 S.Raimondietal./ScientiaHorticulturae219(2017)125–130 maingeneticparametersofasetof290V.viniferacultivars

includ-ingthegenotypesinvestigated:numberofalleles,expected(He) andobserved(Ho)heterozygosity,probabilityofidentity(PI),and polymorphisminformationcontent(PIC).Thetotalnumberof alle-leswas218,andrangedfrom5to16foreachlocus,withanaverage of9.9;expectedheterozygosityvariedbetween28.9%and88.4%, whileobservedheterozygositywasbetween25.9%and92.1%.The probabilityoffindingdifferentgenotypeswiththesameprofileat 22SSRlociappearedtobeverylow(PI=3.23_×10−24):therefore identicalgenotypes(i.e.synonyms)correspondtoidentical pro-files.Thehighdiscriminatingpoweramonggenotypesofthesetof markersusedisalsosupportedbythehighvalueofcumulativePIC (0.710).

Geneticanalysisrevealedthat‘VI-12’and‘VIII-5’(‘Bionda’)are thesamegenotype,asare‘VII-21’(‘S.Martino’)and‘VIII-10’,and ‘XII-26’and‘XII-30’.Thisfindingwasconfirmedbythe morpho-logicalcharacterization.Becauseoftheseduplicates,theunique genotypesstudiedwere39.Thethreecasesofmistakenlabelling couldhaveoccurredeitherduringmaterialpropagationorduring planting/duplicatingofcollections,ratherthaninseedling selec-tion.Unfortunately,thebreeder’snotesonthemorphologyofthe selectedoffspringweretooconcisetoaffordtruevariety identi-fication.Thequestionarisesaboutthecorrect labellingofthese duplicates,i.e.whether,forexample,thegenotypelabelledboth ‘VI-12’and‘VIII-5’actuallycorrespondstothetrue‘VI-12’ortothe true‘VIII-5’(‘Bionda’).Thisproblemwassolvedbycheckingthe consistencyofdeclaredparentages(Table1)withgeneticdata anal-ysis(SSRprofilesinSupplementaryMaterialsandTable3).Inthe exampleofthecontenders‘VI-12’and‘VIII-5’,duetothehigh like-lihoodratioinfavourofthecross‘Bicane’x‘Moscatod’Amburgo’, itcanbepostulatedthatthetruecrossrefersto‘VIII-5’,namely ‘Bionda’.Based onthesamereasoning, thegenotypeconsistent withbeingtheprogenyof‘Chatus’and‘Dolcetto’istheID‘VII-21’, i.e.‘S.Martino’,whilethetruedescendentof‘Furmint’and ‘Malva-siabiancalunga’is‘XII-26’(Table3),ID‘XII-30’beinginconsistent withthisparentage(SSRprofilesinSupplementaryMaterials).

For theother putativeparentages, thelikelihood ratios(LR) calculatedfor proposedparenthoods in relationtoother possi-blecombinationswereallquitehigh,and>theminimumvalue of8.39E+1for‘II-32’(‘Fubiano’)(Table3).Thisindicatesthatthe proposedparentsof‘Fubiano’areabout80timesmorelikelythan otheralternativecombinations.

Thepedigreesof20crossesof39obtainedbyProf.Dalmasso werevalidated(Table3).In13IDs,oneparent(femaleormale)was incorrect,whilefortwoIDsbothparentswereinconsistentwith microsatelliteprofiles.InfourIDsthepedigreecouldnotbeverified forbothparentsbecause,asmentionedabove,oneofthem,namely ‘Pirovano62’,wasmissing.However,moleculardataverifiedthe likelyparenthoodoftheothergenitor,‘MuscatHamburg’.

Inallthewinegrapevarietiesexamined,‘Nebbiolo’was invali-datedasparent,‘Chatus’beingthetruegenotype,enteringthecross eitherasmale orfemalepartner,ashasbeenreported(Torello

Marinonietal.,2009).Sinceone ofthemany Italian synonyms

fortheFrench‘Chatus’is‘Nebbiolo’(popular aroundthevillage ofDronero,hence‘NebbiolodiDronero’),thiscultivarmusthave beenutilisedinthecrossesinsteadofthetrue‘Nebbiolo’.Cipriani etal.(2010)indicated‘Nebbiolo’astheauthenticparentofthetwo IDs‘Cornarea’and‘Nebbiera’;however,thisdemonstratesthatthe accessionof‘Nebbiolo’theyanalysed(maintainedattheCREA-VIT ofConeglianocollectionandprobablythesameusedbyDalmasso) wasmistaken,clearlyactuallycorrespondingto‘Chatus’.

Theproposedparentages(Table1)werenotconﬁrmedforID ‘XIII-11’(‘Cové’)norfor‘XIV-4’,andnorwereotherpossible geni-tors(consistentwiththedescendent’sSSRproﬁles)foundinthe CNR-IPSPgrape geneticdatabase.Thesetwo crosses arein fact

theresultofself-pollinationoftheplantchosenasfemaleparent Table

3 Declared and validated/invalidated parentages of the 39 unique Dalmasso’s interbreeds of wine and table grapes. Likelihood ratios of the proposed families are compared to other possibilities. Probabilities are based on allelic frequencies of 22 n -SSR loci from each genotype and (in brackets) with a 95% upper conﬁdence threshold. X and Y: random cultivar; rel(1): close relative of Parent (1); rel(2): close relative of Parent (2). Accession code Cultivar name Declared parents (names according to VIVC) Parents resulting from genetic analyses Pedigree remarks Parent (1) Parent (2) X x Y (1) x X rel(2) x (1) (2) x X rel(1) x (2) II-26 Vega Furmint × Malvasia istriana Furmint Malvasia istriana 1.29E + 23 1.23E + 13 5.86E + 3 4.91E + 11 2.41E + 3 Validated (2.29E + 19) (1.52E + 11) (2.94E + 3) (7.47E + 9) (1.09E + 3) II-32 Fubiano Furmint × Trebbiano toscano Furmint Trebbiano toscano 3.88E + 12 7.84E + 6 2.36E + 2 8.82E + 7 1.88E + 2 Validated (8.94E + 9) (5.11E + 5) (1.04E + 2) (3.09E + 6) (8.39E + 1) III-3 Muscat Hamburg × Afus Ali Muscat Hamburg Afus Ali 2.75E + 16 8.02E + 7 6.25E + 2 3.18E + 11 2.93E + 3 Validated (4.08E + 13) (5.03E + 6) (2.84E + 2) (7.12E + 9) (1.39E + 3) III-32 Muscat Hamburg × Afus Ali Muscat Hamburg Afus Ali 2.46E + 15 6.56E + 6 4.16E + 2 1.08E + 11 1.50E + 3 Validated (3.89E + 12) (4.37E + 5) (1.75E + 2) (2.46E + 9) (7.21E + 2) III-34 Franca Muscat Hamburg × Afus Ali Muscat Hamburg Afus Ali 1.40E + 14 8.80E + 6 4.51E + 2 5.61E + 9 1.51E + 3 Validated (4.83E + 11) (6.85E + 5) (2.02E + 2) (2.60E + 8) (7.37E + 2) IV-6 Chasselas rose × Csaba Gyoengye Muscat Hamburg Schiava grossa 1.73E + 18 6.40E + 8 1.42E + 3 8.92E + 9 1.76E + 3 Both parents invalidated (2.51E + 15) (3.42E + 7) (6.55E + 2) (2.84E + 8) (7.83E + 2) IV-28 Cornarea Barbera × Nebbiolo Barbera Chatus 1.11E + 14 9.09E + 7 5.35E + 2 2.46E + 9 1.35E + 3 Male parent invalidated a (2.39E + 11) (4.90E + 6) (2.29E + 2) (1.02E + 8) (6.08E + 2) IV-31 Soperga Nebbiolo × Barbera Barbera Chatus 1.86E + 14 8.17E + 7 3.83E + 2 4.94E + 9 1.74E + 3 Female parent invalidated a (3.76E + 11) (4.50E + 6) (1.74E + 2) (2.03E + 8) (7.89E + 2) V-1 Chasselas rose × Csaba Gyoengye Schiava gentile Muscat of Alexandria 2.99E + 15 1.36E + 11 2.49E + 3 1.74E + 8 5.52E + 2 Both parents invalidated (4.86E + 12) (3.80E + 9) (1.17E + 3) (9.87E + 6) (2.63E + 2)

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S. Raimondi et al. / Scientia Horticulturae 219 (2017) 125–130 129

Accessioncode Cultivarname Declaredparents(namesaccordingtoVIVC) Parentsresultingfromgeneticanalyses Pedigreeremarks

Parent(1) Parent(2) XxY (1)xX rel(2)x(1) (2)xX rel(1)x(2)

VI-3 Emilia Bicane×AfusAli Bicane AfusAli 6.88E+14 1.44E+7 3.33E+2 2.22E+10 1.68E+3 Validated

(1.46E+12) (1.02E+6) (1.50E+2) (6.88E+8) (7.96E+2)

VI-6 Bicane×AfusAli Bicane AfusAli 1.38E+16 3.24E+8 4.76E+2 2.08E+10 2.21E+3 Validated

(1.82E+13) (1.33E+7) (2.12E+2) (6.20E+8) (9.95E+2)

VI-9 Bicane×AfusAli Bicane AfusAli 6.64E+13 7.00E+6 3.63E+2 5.92E+9 1.27E+3 Validated

(1.67E+11) (4.73E+5) (1.53E+2) (2.22E+8) (6.08E+2)

VI-24 Bicane×MoscatodiTerracina Bicane Chasselas 3.16E+14 2.26E+8 6.48E+2 2.03E+8 6.75E+2 Maleparentinvalidated

(6.80E+11) (1.08E+7) (2.82E+2) (8.78E+6) (2.91E+2)

VII-21 S.Martino Nebbiolo×Dolcetto Chatus Dolcetto 4.28E+16 2.07E+10 2.39E+3 4.15E+10 2.75E+3 Femaleparentinvalidateda

(6.07E+13) (8.71E+8) (1.16E+3) (1.44E+9) (1.29E+3)

VIII-1 Bicane×MuscatHamburg Bicane MuscatHamburg 7.64E+16 1.82E+9 9.99E+2 3.64E+10 3.16E+3 Validated

(1.18E+14) (7.82E+7) (4.71E+2) (1.31E+9) (1.50E+3)

VIII-5 Bionda Bicane×MuscatHamburg Bicane MuscatHamburg 1.07E+17 4.02E+9 1.40E+3 1.02E+10 1.51E+3 Validated

(1.23E+14) (1.42E+8) (6.26E+2) (3.49E+8) (7.17E+2) IX-2 Schiavagrossa×Muscatof

Alexandria

Schiavagentile MuscatofAlexandria 1.30E+19 9.91E+13 7.98E+3 1.43E+9 8.97E+2 Femaleparentinvalidated (5.64E+15) (1.07E+12) (3.97E+3) (5.84E+7) (4.27E+2)

X-4 Verdiso×Madeleineroyale MadeleineRoyale Verdiso 6.58E+16 9.50E+10 3.34E+3 7.11E+8 1.17E+3 Validated (8.41E+13) (2.83E+9) (1.55E+3) (3.02E+7) (5.03E+2)

X-10 Verdiso×Madeleineroyale MadeleineRoyale Verdiso 7.58E+14 5.63E+9 1.01E+3 8.85E+6 4.79E+2 Validated (1.66E+12) (1.91E+8) (4.71E+2) (6.82E+5) (2.10E+2)

X-12 Sirio Verdiso×Madeleineroyale MadeleineRoyale Verdiso 2.32E+16 4.57E+8 6.09E+2 3.61E+10 2.81E+3 Validated

(3.05E+13) (2.08E+7) (2.74E+2) (1.14E+9) (1.30E+3)

XI-2 MuscatHamburg×AfusAli MuscatHamburg AfusAli 2.03E+16 3.29E+7 5.33E+2 1.20E+11 3.52E+3 Validated (3.64E+13) (2.03E+6) (2.33E+2) (3.54E+9) (1.67E+3)

XI-20 MuscatHamburg×AfusAli MuscatHamburg AfusAli 2.87E+16 1.42E+8 9.32E+2 2.95E+11 4.34E+3 Validated

(4.89E+13) (8.89E+6) (4.23E+2) (8.44E+9) (2.07E+3)

XII-26 Furmint×Malvasiabianca

lunga

Furmint Malvasiabiancalunga 2.02E+24 7.41E+14 6.06E+3 1.52E+13 5.37E+3 Validated (1.24E+20) (4.61E+12) (2.96E+3) (1.42E+11) (2.42E+3)

XII-37 Bussanello Welschriesling×Furmint Welschriesling Furmint 2.11E+20 1.13E+15 1.11E+4 1.51E+10 9.61E+2 Validated

(3.38E+16) (6.61E+12) (5.26E+3) (3.73E+8) (4.32E+2)

XII-40 Welschriesling×Riesling

weiss

Welschriesling Rieslingweiss 1.75E+18 3.73E+7 5.33E+2 2.38E+11 2.20E+3 Validated (9.41E+14) (1.76E+6) (2.12E+2) (3.50E+9) (1.04E+3)

XIII-11 Covè Harslevelue×Malvasiabiancalunga Harslevelue Harslevelue −b _– _– _– _– _Male_parent_invalidated

(self-pollination)a

XIV-4 Grignolino×Sangiovese Grignolino Grignolino – – – – – Maleparentinvalidated

(self-pollination)a

XIV-15 Grignolino×Sangiovese Grignolino Moradella 2.12E+15 5.88E+8 6.35E+2 5.32E+10 3.18E+3 Maleparentinvalidated

(2.95E+12) (2.72E+7) (2.86E+2) (1.48E+9) (1.42E+3)

XV-29 Nebbiera Nebbiolo×Barbera Chatus Barbera 6.29E+15 1.22E+10 1.60E+3 1.01E+9 7.41E+2 Femaleparentinvalidateda

(7.89E+12) (4.02E+8) (7.25E+2) (4.02E+7) (3.25E+2)

XV-31 Albarossa Nebbiolo×Barbera Chatus Barbera 2.35E+17 4.36E+8 7.41E+2 1.57E+12 4.94E+3 Femaleparentinvalidateda

(1.83E+14) (1.87E+7) (3.19E+2) (2.97E+10) (2.34E+3)

XV-34 S.Michele Nebbiolo×Barbera Chatus Barbera 3.54E+12 1.56E+8 4.96E+2 1.24E+7 2.41E+2 Femaleparentinvalidateda

(1.13E+10) (7.79E+6) (2.22E+2) (7.99E+5) (1.09E+2)

XVI-8 Valentino Nebbiolo×Dolcetto Chatus Dolcetto 5.46E+17 3.41E+9 1.29E+3 4.06E+12 6.19E+3 Femaleparentinvalidateda

(4.49E+14) (1.38E+8) (5.85E+2) (7.74E+10) (2.97E+3)

XVII-25 Passau Dolcetto×Nebbiolo Dolcetto Chatus 2.31E+18 3.00E+13 7.94E+3 2.63E+9 1.23E+3 Maleparentinvalidateda

(1.31E+15) (3.60E+11) (3.71E+3) (1.14E+8) (5.59E+2)

XVIII-3 Viola MuscatHamburg×Pirovano62 MuscatHamburg – unveriﬁable

XVIII-12 Liana MuscatHamburg×Pirovano62 MuscatHamburg – unveriﬁable

XVIII-21 Giovanna MuscatHamburg×Pirovano62 MuscatHamburg – unveriﬁable

XVIII-24 Teresita MuscatHamburg×Pirovano62 MuscatHamburg – unveriﬁable

a_{Conﬁrmation}_of_prior_publication.

(6)

130 S.Raimondietal./ScientiaHorticulturae219(2017)125–130 (asreportedbyCiprianietal.,2010for‘Cové’),likelybecauseofa

mistakeintheemasculationprocess.Inbothcrosses,but particu-larlyinthesecondone,theplant’svigourwasdepressed,andleaf morphologicalanomaliesaroseattimes.

In ‘VI-24’ and ‘XIV-15’, thepollen donor wasfound tobe a differentcultivarfromthatexpected,likelybecauseofpollen con-tamination.Asinthecaseof‘Chatus’beingmistakenfor‘Nebbiolo’, inone cross (‘IX-2’)the trueparentwas‘Schiava gentile’(alias ‘Urbanblau’),alesswidespreadcultivarthanthedeclaredgenitor ‘Schiavagrossa’,knownas‘Trollinger’and‘Frankenthal’inother centralEuropeancountries,andnamed‘Schiavone’inDalmasso’s notes.

Fortwotablegrapevines(‘IV-6’and‘V-1’),bothparentswere invalidated:thismightbeduetothemistakenidentityofboth par-entsor,morelikely,toamistakeinseedlinglabelling,orevenin materialpropagationfrommotherplants duringﬁeldcollection establishmentorduplication.Thelackofadetailedmorphological descriptionfromthebreederpreventstrue-to-typeveriﬁcationof seedlings.

As already mentioned, the descent of the four offspring of ‘Pirovano62couldnotbeingveriﬁed,since‘Pirovano62’appears tohavedisappearedfromEuropeancollections.InthelargeINRA collection at Domaine de Vassal (France), where ‘Pirovano 62’ shouldbemaintainedaccordingtotheVitisInternationalVariety Catalogue(www.vivc.de),theconservedaccessionprobably corre-spondsinsteadto‘Pirovano61’(asiblingof‘Pirovano62’),asINRA scientistssuspect(J.-M.Boursiquot,personalcommunication).

Veriﬁcationofparentages declaredby breeders,throughthe inheritanceofSSRmarkers,revealed43%ofinvalidatedpedigrees intheinvestigatedcross-breeds(15outof35veriﬁedpedigrees). Thispercentageishighcomparedtothosereportedinother stud-ies:Lacombeetal.(2013)analysed381moderncultivars,andfound breeder’sdatatobeinvalidin126genotypes(33%).Inthepresent study,theuseof‘Chatus’insteadof‘Nebbiolo’inasubstantial num-berofIDshad amultipliereffectontheinaccuracyofbreeder’s data.Vargasetal.(2009)alsofoundasimilarpercentageof incor-rectpedigrees(42%,13incorrectpedigreesversus18correct,as reportedintheliterature).

4. Conclusions

Alongsideinvalidatedcrosses,likelyduetopollen contamina-tion(whenthemaleparentwasincorrect),severalcasesofcultivar mislabelling(eitherwhenbothparentswereinvalidatedorwhen thesame genotypewas labelleddifferently) werefoundin the varietiesinvestigated.Thisindicatesthatfurthermislabelling dur-ingvegetativepropagationtoestablish/duplicatecollectionsmay havebeenaddedtopossiblecross-breedingerrors,makingvarietal identiﬁcationessentialfornewly-bredvarieties.

Theeconomicexploitationofnewbreedsbygrapefarmersand thewineindustrymaybeconsiderablyaffectedbytheircorrect identiﬁcationandparentage.Astobreeder’sactivity,in particu-larrelated totablegrapes,thecorrect information providedon thegeneticbasis ofthecrosses obtainedmaycontributetothe developmentoffurtherbreedingprograms.Lastly,theparentage veriﬁcationand genetic characterizationof 39 grapevine cross-breedsobtainedbyProf.Dalmassohasprovidedadditionalinsight intotheavailablegrapevinediversity.

Funding

ThisresearchwasfundedbytheAgriculturalResearch Founda-tion(AGER,Italy)[grantnumber2010-2104].

Acknowledgments

TheauthorsthanktheAgriculturalResearchFoundation(AGER, Italy) for ﬁnancial support and Giovanni Dalmasso Foundation forpatronage.WearegratefultoMassimoGardimanofthe Con-siglioperlaRicercainAgricoltura(CREA-VIT),Conegliano(Italy) forproviding two samplesfromtheircollections.We gratefully acknowledgetheassistancegivenbyJ-M.Boursiquot(INRA, Mont-pellier, and Domaine de Vassal, France) who provided useful informationon‘Pirovano61’and‘Pirovano62’.

AppendixA. Supplementarydata

Supplementarydataassociatedwiththisarticlecanbefound, intheonlineversion,athttp://dx.doi.org/10.1016/j.scienta.2017. 02.044.

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