Recent Achievements and Current Status of Vitis vinifera Germplasm in South East Europe

Maja Žulj Mihaljević1, Darko Preiner1, Silvio Šimon1, Klime Beleski2, Frida Carka3, Gheorghe Savin4,

Mihai Botu5, Ivan Pejić1 1)University of Zagreb, Faculty of Agriculture, 10000 Zagreb, Svetošimunska 25, Croatia (mzulj@agr.hr); 2) Institute of Agriculture, Dept. for Viticulture and Enology, Aleksandar Makedonski bb, 1000 Skopje, Republic of Macedonia 3) Agricultural University of Tirana, Genetic Resources Center, Tirana, Albania 4) Research and Practical Institute for Horticulture and Food Technologies, Dept. of Genetic Resources and Amelioration 59, Vieru Str., MD-2070, Chisinau, Moldova 5) University of Craiova, Faculty of Agriculture and Horticulture, Dept. of Horticulture and Food Science, 13, A. I. Cuza St., 200585 Craiova, Romania

Ampelographic and molecular characterization of national germplasm collections and local varieties has been done in many European countries, but in countries of South East Europe, grapevine material is still not properly genetically evaluated and compared due to fragmented projects and approaches. Even if the genotyping using microsatellite (SSR) markers was done, results were not standardized, stored and compared with international databases, so origin of some material is still unknown/misunderstood. Well managed genetic grapevine resources provide basic material for selection and improvement trough breeding, but also can provide a starting point for revitalization of rare varieties. Recently, several independent projects of regional collaboration for grapevine germplasm analysis made a thorough inventory of native South East European germplasm. Examples of those are projects like „Identification, characterization and conservation of old and autochthonous wine varieties in South Eastern European countries“ which included eight different partners from South East Europe as part of SEEDNet projects and whose results were recently published (Žulj Mihaljević et al., 2013); project “Preservation and establishment of true-to-type and virus free material of endangered grapevine cultivars in Croatia and Montenegro”, as part of SEE-ERA.net projects or GrapeGen06 (GG06) project "Management & Conservation of Grapevine Genetic Resources„ which included 25 participants, amongst which were also some South Eastern European countries like Croatia, Greece and Moldova and whose results are available through European (EU) Vitis Database (Maul et al. 2012). It contains SSR data for more than 4000 different Vitis accessions. Aim of this study was to give an overview of the current status of Vitis vinifera germplasm in neighboring countries of South East Europe through survey of recent literature and projects of regional collaboration.

Conclusions Acknowledgements: This research is the outcome of several research projects: Seednet, GrapeGen 06, and SEE-ERA.net plus (ERA91), for details see Introduction chapter.

Results and current status of national germplasms are described per country (Table1). Albania Genetic and ampelographic data of Albanian accessions remain unavailable in EU Vitis Database, although there was an initiative to genotype autochthonous accessions (Maul et al. 2003). We compared Albanian accessions from Seednet project with work of Ladoukakis et al. in 2005, first attempt of ascertaining genetic identity of Albanian cultivars. Only two accession names appear in both studies: ′Sheshi i bardhe′ and ′Serina e zeze′. Visual comparison of common loci in both studies indicates that these accessions which share the same name don’t have same SSR profile. Since ′Sheshi i bardhe′ is one of the most spread varieties in Albania, further ampelographic work is needed. Bosnia and Herzegovina Only recently genetic characterization of native B&H germplasm has been done (Tomić et al. 2012, Leko et al. 2012), and this late start is partly caused by the fact that this country is still politically divided and that there’s no collection vineyards. Recent projects addressed this issue and results from Table1, like the fact that supposedly autochthonous variety ′Dalmatinka′ is in fact ′Alicante Bouschet′, one of the most cultivated red varieties in B&H, will help in establishment of autochthonous germplasm collection. Croatia Data for more than 100 Croatian accessions is stored in EU Vitis database (Picture 1). Croatian variety ′Kamenina′ matches with ′Corredera′, which is known to be a ′Palomino′ X ′Cardinal′ cross. Accessions ′Medna bijela′ and ′Bumba bijela′ are the same as Herzegovinian accession ′Zložder′. This is not surprising, since they originate from neighboring regions that were always economically connected. ′Stara belina′ matched with Romanian variety ′Slaviţă′ (′Slavitza′). Its origin remains unknown since its Romanian name suggests its Slavic origin. It has been documented as a pre-phyloxeric variety before 1900’s in south west Romania and ampelographers opinion is that it can be a local variety or that has been introduced long time ago. An unkonwn accession from Moslavina region matched Greek accession ′Pamidi ′, Bulgarian variety ′Pamid′ and is also traditionally grown in Serbia, under the name ′Plovdina′, from where it probably came to Croatia. Macedonia Only two accessions are listed in EU Vitis database. Comparing 14 accessions genotyped in Seednet project with Štajner et al. (2009), we found that all four common accessions (′Vranec′, ′Kratoshija′, ′Stanushina′, ′Crven drenok′) match on three common SSR loci and can be considered true to type. Accession ′Bojanka′ is the same as ′Grand noir′, coming from a cross of ′Aramon′ x ′Petit Bouschet′ so we can consider it as synonym for ′Grand noir′. Accession ′Kratoshija′, which is widely grown in Macedonia and spread from Montenegro, matches genotypes of ′Zinfandel′/′Crljenak kaštelanski′/′Primitivo′/′Kratošija′. Moldova Currently, in EU Vitis database, certain level of information for 421 Moldavian accessions are stored, but no SSR marker data. Four accessions in Seednet project have not been mentioned and two of them have its match in EU Vitis. Accession ′Sghihara′ is the same as genotype of ′Plavay′, whose origin is in fact from Moldova. ′Tamaioasa′, appears to be local synonym of ′Muscat Blanc à Petits Grains′. Ghetea et al. (2012) claim that ′Tămâioasă Românească′ is a very old and autochthonous Romanian cultivar, known in ancient Greece as ′Anathelion moschaton′, during Roman Empire as ′Aspinae′ and that is cultivated in Romanian vineyards from ancient times. ′Tămâioasă Românească′ and ′Tămâioasa′, Moldavian accession, are the same genotypes. ′Muscat Blanc à Petits Grains′ is an ancient cultivar and can be found under different synonyms in many wine growing regions.

Romania Currently, no SSR data is stored in EU Vitis database on the Romanian varieties although some genotyping studies have been conducted (Coste et al., 2010; Gheţea et al. 2010; Gheorghe et al., 2010). Synonyms were found for six supposedly autochthonous Romanian varieties, as listed in Table 1. ′Ţȃţa caprei alba′ matches with three accessions: German ′Cornichon blanc′, Greek ′Tsougiannides′ and Serbian accession ′Ćilibarika′, table grape cultivar, whose origin is considered to be Middle East. Accession ′Gordan′ turns out to be synonym for ′Furmint,′ variety used for famous Tokay wine which also has a synonym in Croatia.

Figure 1. Proper managment of autochthonous grapevine collections is enhanced by good insight about number of synonyms, homonyms and curation errors through SSR genotyping.

Comparison of internal SSR grapevine database and thorough survey of the literature revealed many synonyms or curation errors across different countries of South East Europe. Detected synonyms between neighboring countries were mostly unknown before, but since South East European countries share common history, certain level of admixture between cultivars can be expected. Results show necessity of shared database and use of reference varieties and common set of microsatellite markers. This results should also be a lead to ampelographers to compare their ampelographic records and make further effort to properly manage their germplasm collections.

Introduction

Recent achievements in SSR genotyping of some supposedly autochthonous and unknown accessions will be presented through their comparison with internal microsatellite database (University of Zagreb) built up from recent projects, the European Vitis Database and publications dealing with genotyping of grapevine resources of South East European countries in order to determine synonyms and/or homonyms or possible naming errors. Highly polymorphic microsatellite loci used for construction of EU Vitis database were proposed as „standard set” for grapevine analysis. In above mentioned projects, the same SSR loci were used for genotyping, thus enabling comparison of results. Genetic profiles were compared either with Excel Microsatellite Toolkit software (Park, 2001) or by visual inspection when comparing profiles from publications. Later included either comparing allele values with the help of reference variety or comparison of absolute values among alleles and homozygote/heterozygote state of locus.

Country of Analyzed accession Compared with (source of SSR data, country of accession's origin) Comment accession's origin: (source of SSR data):

Albania Sheshi i bardhe (Seednet) Sheshi i bardhe (Ladoukakis et al., 2005, Albania) SSR data don't match Serina e zeze (Seednet) Serina e zeze (Ladoukakis et al., 2005, Albania) SSR data don't match Bosnia and Kadarun bijeli (Seednet) Apsimo prosotsanis (European Vitis Database, Greece) synonym Herzegovina Dalmatinka (Seednet) Alicante Bouschet (European Vitis Database, France) synonym

Plavka (Seednet) Plavina (European Vitis Database, Croatia) synonym Plavka (Tomić et al. 2012, B&H) synonym Croatia Kamenina(Seednet) Corredera (European Vitis Database, Germany) synonym

Okatica vrgorska /Bljuzgavac (internal database) Zložder (internal database, Herzegovina) synonym HR7 (SEE-ERA.net) Pamidi (European Vitis Database, Greece); synonym

Pamid (Hvarleva et al. 2004, Bulgaria); synonym Plovdina (Bešlić et al. 2012, Serbia) synonym

Stara belina (SEE-ERA.net) ′Slaviţă′ (Seednet, Romania) synonym Macedonia Crven valandovski drenok (Seednet) Malvasia rei (European Vitis Database, Italy) misnaming

Kratoshija(Seednet) Primitivo (European Vitis Database, Italy) synonym Crljenak kaštelanski (European Vitis Database, Croatia)

Kratošija (SEE-ERA.net, Montenegro) synonym synonym

Bojanka(Seednet) Grand noir (European Vitis Database, Germany) synonym Moldavia Sghihara(Seednet) Plavay (European Vitis Database, Germany) synonym

Tamaioasa(Seednet) Muscat d'Alsace rose (European Vitis Database, France) synonym Romania Coarnă neagră (Seednet) Lattuario nero (European Vitis Database, Italy) synonym

Ţȃţa caprei albă (Seednet) Cornichon blanc (European Vitis Database, Germany); misnaming Tsougiannides (European Vitis Database, Greece); synonym

Ćilibarika (Bešlić et al., 2012, Serbia) synonym Gordan (Seednet) Furmint (European Vitis Database, Italy); synonym

Moslavac(European Vitis Database, Croatia) synonym Teişor (Seednet) Semillon (European Vitis Database, France) synonym

Braghină albă (Seednet) Kopcak (European Vitis Database, Czech Republic) Kopcak not true to type Negru moale (Seednet) Cot (European Vitis Database, France) synonym

Materials and Methods

Results

Table 1. List of synonyms and possible misnaming