Project ADER: 3.1.3.

Development and modernization of grapevine

germplasm collections aiming to long-term

preservation of the national value plant material

Project coordinator:

NATIONAL RESEARCH and DEVELOPMENT INSTITUTE

for BIOTECHNOLOGY in HORTICULTURE

ŞTEFĂNEŞTI – ARGEŞ

Project manager : Ph. ing. Constantin Tănăsescu

Partners in the consortium

P1 Research and Development Institute for Viticulture and Oenology Valea Călugărească

P2 Research and Development Station for Viticulture and Oenology Blaj

P3 Research and Development Station for Viticulture and Oenology Bujoru

P4 Research and Development Station for Viticulture and Oenology Drăgăşani

P5 Research and Development Station for Viticulture and Oenology Odobeşti

P6 Research and Development Station for Plant Culture on Sandy Soils Dăbuleni

Total Budget 712.850 lei / co-funding -Accomplished 708.066 lei / co-funding -

General information

• The networking of research units involved in conservation andexploitation of the grapevine genetic resources is essential.Teamwork ensures the enhancement of human experience as well asthe correct assessment and valorization of the existing grapevinebiodiversity in germplasm collections.

• The ADER 3.1.3 project, had the following objectives:

• I. The assessing of present situation of the grapevine germplasmcollections;

• II. Characterization of the assortment of collections, especially of theold cultivars, considered autochthonous, applying standardized andupdated methods for ampelographic descriptors, in parallel with eno-carpological, biochemical and molecular ones;

• III. Enlarging and upgrading of vine germplasm collections accordingto the OIV (International Vine and Wine Organization) standards.

ResultsI. The assessing of present situation of grapevine germplasm collections

The objectiv is: Updating the records of germplasm collections by a unique system and in

accordance to the international requirements. Centralization information regarding the

assortment from vine collections held by the participating units

Item Total number of cultivars / cultivars from other countries Total no. / % of autochthonous cvs

2015 / 2018CP P1 P3 P4 P2 P5 P6

Cvs. and clones -

Seedless table

16/11

16/11

57/53

(+1) 58/53

6/3

6/3

25/22

25/22

-/- 3/1

3/1

5/2

5/2

2015= 112 / 17,8 % (20)

2018= 113 (+1) / 17,7 % (20)

Cvs. and clones –

Seeded table

68/16

(+5) 73/16

575/533

(+26)601/533

30/14

(+19) 49/14

97/71

97/71

26/7

28 (+2)/7

33/15

33/15

16/7

17/8 (+1)

2015= 845 / 21,5 % (182)

2018= 898(+53) / 26 % (234)

Cvs. and clones -

White wines

96/4

(+2) 98/4

395/344

(+23)418/344

39/23

(+2)41/23

68/22

68/22

55/23

24(-31)/16(-7)

63/30

64(+1)/31

12/5

12/5

2015= 728 / 38 % (277)

2018= 725(-3) / 38,6 % (280)

Cvs. and clones -Red and rose wines

59/10

(+2) 61/10

242/222

(+18)261/222

28/24

28/24

50/28

50/28

12/8

3 (-9)/1 (-7)

37/21

38 (+1)/21

14/3

15/4 (+1)

2015= 442 / 28,5 % (126)

2018= 456(+13) / 32 % (146)

Cvs. and clones –

Flavour wines

13/8

(+2) 15/8

27/22

27/22

7/2

9/4 (+2)

9/5

9/5

12/6

2(-10)/2(-4)

15/10

15/10

4/2

4/2

2015= 87 / 36,8 % (32)

2018= 81 (-6) / 34,6 % (28)

Cvs. and clones

rootstocks

24/14

24/14

35/30

35/30

--

- -

47/41

47/41

4/2

4/2

--

- -

-/-

- -

2015= 110 / 20,9% (23)

2018= 110 / 20,9% (23)

Inter-specific

hybrids

27/17

27/17

85/77

85/77

2/1

2/1

--

--

24/20

8(-16)/4(-16)

78/76

51 -27)/49

1/-

1/-

2015= 217 / 11,9% (26)

2018=174 (-43) / 14,9 % (26)

Species / wild

populations

9/-

9/-

8/8

8/8

--

--

13/13

13/13

-/-

-/-

-/-

-/-

-/-

-/-

2015= 30 / 30% (9)

2018= 30 / 30% (9)

TOTAL cvs. /

autochthonous

cvs.

312 / 232

323(+11) /243

1424 / 135

1492 (+68)/203

112 / 45

(+23)135/66

309 / 107

309/ 107

133/67

69 (-64)/37

229/76

204(-25)/77

52/33

54 (+2)/33

2015= 2571 / 27 % (695)

2018=2586(+15)/29,6 % (766)

ResultsI. The assessing of present situation of grapevine germplasm collections

Total number of varieties in collections and number of native varieties,

compared 2015 and 2018 (Total in the 7 research units)

0

100

200

300

400

500

600

700

800

900

Total Autohtone Total Autohtone Total Autohtone Total Autohtone Total Autohtone Total Autohtone Total Autohtone

112

20

845

182

728

277

442

126 87

32

110

23

217

26

113

20

898

234

725

280

456

146

8128

110

23

174

26

2015 2018

Table seedless Table seeded White wines Red and rose wines Flavour wines Rootstocks Inter-specific hybrids

The proportion of accessions deemed as autochthonous

(% of total accessions of the same type) between 2015-2018

0

5

10

15

20

25

30

35

40

17.8

21.5

38

28.5

36.8

20.9

11.9

30

17.7

26

38.6

32

34.6

20.9

14.9

30

2015 2018Table seedless Table seeded White wines Red and rose wines Flavour wines Rootstocks Hybrids Wild vines

The varieties considered as autochthonous (766), important for the respective

wine-growing areas, have been identified and have to be fully characterized

(by ampelography and molecular methods)

ResultsI. The assessing of present situation of grapevine germplasm collections

Results

I. The assessing of present situation of grapevine germplasm collections

Number of cultivars, clones and hybrids of Vitis genus,

existing only in a single germplasm collection

Grapevine

cultivars PC P1 P2 P3 P4 P5 P6 Total

Cvs. and clones -

Seedless table 9 19 - - 17 - - 45

Cvs. and clones –

Seeded table 1 313 6 - 25 4 - 349

Cvs. and clones -

White wines 1 353 12 13 4 - 383

Cvs. and clones -

Red and rose wines2 104 9 7 17 5 - 144

Cvs. and clones –

Flavour wines - - 3 - 3 - - 6

Interspecific hybrids

and rootstocks 1 42 12 - 1 - - 56

Total – varieties

existing in a single

collection

14 831 42 7 76 13 - 983

Were identified the varieties present in one single germplasm collection; for these

accessions are necessary protection and propagation measures aiming to reduce

the risk of their disappearance

II. Characterization of the assortment of collections, especially of the old cultivars,

considered autochthonous, applying standardized methods

• In all units participating in the project, had applied the same methodology for the ampelographic characterization to 4 reference varieties (internationally recognized varieties and present in all collections) and to a number of varieties of vines considered indigenous and representative for the respective wine-growing zone.

• Update of the selected assortment: 75 old cultivars of Vitis vinifera subsp. vinifera, considered as autochthonous (24 table grapes, 34 cvs. for white wines, 13 cvs. for red wines, 4 cvs. for flavour wines) and 8 varieties of Vitis vinifera subsp. sylvestris.

• This plant material offers a wide range of properties and favourable characters, expressed through valuable agro-biological and economical phenotypes, constituting a genetic material useful as propagation material and as genes source for grapevine breeding (obtaining of new varieties or clones).

• To verify the authenticity of the accessions present in each collection were applied the same procedures and numerical evaluations, based on the same system, international recognized – "OIV descriptor list for grape varieties and Vitis species (2nd edition)" (2009).

For the first time, ampelography descriptions were presented on 8 varieties of

wild vines from 3 different wild areas (Hrinova-Cazane, Greaca and Letea Forest)

OIV-001 opening of the shoot tip

Vs10 G Vs Letea 1 Vs3eb G

OIV-051 color of young leaf

Vs3eb G Vs Letea 1 Vs9dm H

Mature leaf: size (065), shape (067),

number of lobes (068), etc.

Vs6 S Vs9 G Vs9dm H

Vs12 G Vs14 G Vs Letea 2

Vs3eb G Vs14 G

Vs10 G Vs Letea 2

Bunch: size (202), compactness (204),

color of skin (225), etc.

Ampelography description of the selected cultivars

GORDAN (ROM051-257)

Descriptors related to trueness to type:

Trueness to type (of the variety) yes

“Remarks to the accession name” true name

“Variety number VIVC” 5544

“Confirmation by ampelography” yes

“Confirmation by SSR-markers” yes

“Confirmation by bibliography” yes

Synonyms: 60 (Gorgan, Gordan alb, Corb alb, Băldoaie, Gordan mic,Gordan mare, Gordană, Timpurie, Giordan, Iordovana, Jordan, Iordan,Jârdovâny, járdovány fehér, etc.)

Ampelography description of the selected cultivars

NEGRU VARTOS (ROM051-263)

Descriptors related to trueness to type:

Trueness to type (of the variety) yes

“Remarks to the accession name” true name

“Variety number VIVC” 8466

“Confirmation by ampelography” yes

“Confirmation by SSR-markers” yes

“Confirmation by bibliography” yes

Synonyms: 20 (Corb, Negru bătut, Gordan negru, Beilar Cherasi,Negru bulgar, Negru tare, Negru tare, Negru bulgăresc, Gordan negru,Cerno tvtrdo, Garvan, etc.)

Molecular analysis

Complete characterization of the accessions means:

description with standardized descriptors +

molecular analysis with a set of at least nine SSR markers

Were analysed 54 grapevine cultivars, considered autochthonous;

DNA extraction - Qiagen DNeasy Plant mini-kit

9 SSR markers: VVS2, VVMD5, VVMD7, VVMD25, VVMD27, VVMD28,

VVMD32, VrZAG62, VrZAG79

The data obtained were compared to the existing databases from :

- Centre of Viticultural Research (CREA-VIT), CONEGLIANO, Italia / Manna

Crespan;

- JKI - Julius Kühn-Institut für Rebenzüchtung GEILWEILERHOF, Germania /

Erika Maul;

- INRA, Montpellier - VASSAL collection, Franţa / Thierry Lacombe şi Valerie

Laucou

The results of the molecular analyses were certified

and have been added to the international databases

For 11 cultivars were confirmed the synonyms mentioned in old reference

documents: Ardeleanca with Bakar belyi; Creață with Creaţă de Banat and Kreaca;

Bacator with Bakator roz; Braghină roz with Braghină roșie rară; Cadarcă with Kadarka

Kek; Ceauș alb with Chaouch blanc; Majarcă albă with Slankamenka bela; Negru vârtos

with Mavrud Varnenskii; Plăvaie with Plavay; Tâța caprei albă with Tsitska kaprei.

For 3 old cultivars were not confirmed the synonyms:

- Busuioacā de Bohotin proved to be somatic mutant of Muscat à petits grains

blancs (Greece) for berry colour and it is not synonym with Muscat rouge de Madere

(Constantinescu et al.1960);

- Galbenā uriaşā is a different cultivar and not a selection of Galbenā de Odobeşti

(NICOLEANU 1900 ), it is synonym with Mirkovaca (Croatia);

- Tămâioasă românească proved to be a descendant of Muscat à petits grains

blancs (France) and it is not a synonym with this.

Were identified new synonyms for 5 accessions about were not found information

in the old documents:

Braghină albă synonym with Dinka zoeld (Hungary);

Galbenă măruntă synonym with Kakotrygis (Greece);

Ţâţa caprei neagră synonym with Hora (Bulgaria);

Ţâţa vacii albă synonym with Halholyag (Ukraine);

Ţâţa vacii neagră synonym with Kozi Cici cherveni (Bulgaria).

The results of the molecular analysis were certified and have been added to the international databases.

Were documented new synonyms for 4 analysed accessions:

Ceauș roz with Chaouch rozovyi (Turkey); Coada oilor with Juhfark (Hungary);

Țâța vacii albă with Halholyag (Ukraine); Cârcioasă with Balint weiss (Hungary).

For the first time were proved the synonymies among:

- Gordan = Iordană = Zemoasă;

- Galbena de Odobeşti = Zghihară de Huşi;

- Bătută neagră = Negru românesc

The genetic profiles obtained for the 3 accessions Moroștină, Negru mare and Românie

proved their unicity in the European grapevine germplasm collections

15 cultivars revealed identical profile with those already registered in European Vitis

databases.

For 4 varieties, differences were found from the descriptions in the reference literature and

their SSR profiles did not matched with any indicated variety as possible synonyms.

Morphologic description with standardized descriptors (OIV) and molecular methods

with SSR markers, used together, have proven to be effective for:

confirmation of the authenticity of the biological material, identification of unique

genotypes, the synonyms cultivars and the incorrect registered ones.

ResultsIII. Enlarging and upgrading of vine germplasm collections according to

the OIV (International Vine and Wine Organization) standards

Adopt and implement a unitary system for presenting their owngermplasm collections, to describe and characterize the accessions.Standardization of the used forms for:

a) registration of the existing and / or introduced cultivars;b) the most complete characterization of the accessions from the own germplasm collections of the units participating in the project;c) documenting the cultivars with the purpose of exploiting theplant material to different beneficiaries.

Based on the full description and compliant with the requirements for European germplasm collections, the description files for the cultivars held in each collection were completed.

The correct identification of the cultivars is important for curators, viticulturists, winemakers and consumers; it is also a proof of a fair traceability system throughout the process of obtaining planting material, grape production, winemaking and marketing.

Results

III. Enlarging and upgrading of vine germplasm collections according to

the OIV (International Vine and Wine Organization) standards

Ex situ germplasm collection established by PC

Category/type No. varieties Planted in

2017

Rest for

planting

Cultivars for white wines 59 42 17Cultivars for flavour wines 6 5 1Cultivars for red wines 31 26 5Cultivars for table seeded 42 28 14Cultivars for table seedless 11 6 5Rootstocks 11 9 2V. sylvestris 11 8 3TOTAL 171 124 47

From which:

- 65 cultivars (considered autochthonous) were received from Faculty of

Horticulture Bucharest and from RDSVO Drăgăşani- 9 cultivars were received by exchanging plant material with partners in

consortium (RDIVO Valea Călugărească - P1, RDSVO Odobeşti - P5)

RESULTS

Dissemination of the results obtained within the project

Scientific papers published in the country 16

Scientific papers published abroad 2

Posters and scientific papers presented at national scientific events

with international participation 12

Scientific events organized by PC and P4 (RDSVO Drăgăşani) 2

It was published the book entitle "Strategies and principles forconservation of grapevine genetic resources in ex situ collections"Editure University of Piteşti, ISBN 978-606-560-603-6

Information through the mass-media: P1, P3 and P4 participated inagricultural TV shows where they presented some of their own results, as wellas shortcomings in the maintenance of vineyards and harvesting the grapeyield: TVR 1, Pro TV - national stations, Valcea 1 and TVR 3 Craiova - localTV stations.

Dissemination of the results obtained within the project

ADER 3.1.3.

NRDIBH Ștefănești-Argeș - internal session entitled: “Grapevine genetic

resources - germplasm collections, complete characterization, capitalization”

- 8 June 2017

Dissemination of the results obtained within the projectADER 3.1.3.

Round table – ”Saving the national vineyard patrimony"– RDSVO Dragasani”

19 April 2018

Dissemination of the results obtained within the project

ADER 3.1.3.

Scientific documents presented and published

Plant material in ex situ germplant collection - PC