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Ampelographic and DNA Characterization of Local Grapevine Accessions of the Tuscia Area (Latium, Italy)

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Ampelographic and DNA Characterization of Local Grapevine Accessions of the Tuscia Area (Latium, Italy) Research Note Ampelographic and DNA Characterization of Local Grapevine Accessions of the Tuscia Area (Latium, Italy) Massimo Muganu , l * Gerald Dangl,2 Maui Aradhya,3 Manuela Frediani,4 Angela Scossa, 1 and Ed Stover' Abstract: The presence of local vines in the Tuscia area (Latium region, Italy) was documented by historical sources. Ampelographic and molecular characterization (35 morphological descriptors and 16 microsatellite loci) was carried out on accessions belonging to the most frequently mentioned local vines. SSR markers have enabled us to distinguish cases of local synonymy and microsatellite profiles have permitted comparison with previously reported profiles indicating synonymy and divergence with vines in other collections. Ampelograpliic traits showed that some accessions of the same genotype differ in minor characteristics that can have a role in clonal evaluation. Key words: Vitis vinifera, germplasm, SSRs, grape quality The Latium region, situated in the center of Italy. has graphic studies described several vines that contained the a varied grape germplasm and a winemaking tradition name Greco (Bullettino ampelografico 1881). In the Mid- that predates the Roman civilization. In the Tuscia area dle Ages, greco was an adjective that referred to sweet of Latium (province of Viterbo), seeds of the cultivated and long-lived wine, with high alcohol, and well-known grape Vitis vinijera saliva date to the 10th century ac, in Tuseia (Lanconelli 1994). In the 19th century, Pagadeb- testifying to the antiquity of grape consumption and pro- ito (also called Rornanesco and Scassadehito in Tuscia) duction (Delpino 2005). Wines produced in the Tuscia was one of the most cultivated varieties in the area, very area were popular and well-known during the Middle productive, and yielding wine in quantity (Cinelli 1884). Ages and the Renaissance (Baeei 1596). However, over Moscato was the variety used for the production of the the past century, the land used for viticulture in the Tus- most important wine of Tuscia during the Middle Ages cia area and the quality of its wines have declined, partly and Renaissance (Bacci 1596). because of decreased interest in local varieties. Conse- The preservation of gerinplasm involves many steps, quently, there is a risk that gerrnplasm with potential en- and the aim of this study was to obtain molecular and vironmental adaptations may be lost altogether (Natali ampelographic characterization of accessions belonging and Bignami 1988, Muganu et al. 2007). to local vines of the Tuscia area, to identify the genetic At present there is a campaign to revitalize wine pro- profile and morphologic characteristics of local vines, duction in Tuscia, beginning with the recovery of local and to clarify the presence of synonymies with other varieties, such as Cannaiola di Marta, Rossetto, Greco vines of the region. Pagadebito, and Moscato. Cannaiola di Marta (Cinelli 1884) is a vine used in table wine and presently grown in Materials and Methods Tuscia in limited areas. The historical importance of Ros- There has been a concerted effort to recover biotypes setto in the Tuscia has been documented (Gallesio 1833) of vines of the Tuscia area since 1987. The material has and it is described as one of the most cultivated vines, been cataloged as to town of origin, year, details of grow- late harvested, less susceptible to fungal pathogens, and ing location (private farms, old vineyards, vegetable gar- producing a long-lived wine (Cinelli 1884). Early ampclo- dens, gardens, single plants), and accession name supplied by the donor. In 1995 the different accessions were used to establish the DIPROV (Dipartimento di Produzione Ve- Dipartimento di Produzionc \'cgetalc, Universitâ della Tuscia, via S. Camillo de Lellis snc, 01100 Viterbo, Italy; 217oundation Plant Services. University getale) grape germplasm collection at the University of of California, Davis, CA 95616; 3USDA, ARS National Clonal Germplasm Tuscia experimental farm in Viterbo. Preliminary histori- Repository, University of California, Davis, CA 95616; and 4 Dipartirnento di cal and bibliographic research was conducted to document Agrobiologia ed Agrochimica, Università della Tuscia, 01100 Viterbo. Italy. the historical presence of the vine varieties in the area. *Corresponding author (email: [email protected]; tel: 39 0761 357325; fax: 0761 357531) Sampling and plant material. All ampelographic and Manuscript submitted May 2008, revised Sept 2008, accepted Dec 2008. phenological data and samples designated as DIPROV and Publication Costs of this article defrayed in part by page fees. that were used for DNA extraction were collected from Copyright L 2009 by the American Society for Etiology and Viticulture. All plants grown in the collection of the University of Tus- rights reserved. cia (]at. 42°25'21'N; long. 12°04'45'E). The vines were 110 Am. J. Enol. Vitic. 60:1 (2009) Characterization of Tuscia Grapevine Accessions - 111 (Qiagen, Valencia, CA) following the manufacturer's grafted to 420A, Guyot-trained, and spaced 3 m x 1.5 in. protocol. Each accession was sampled twice. PCR am- All plants were subject to the same climatic and agro- plifications were performed on a GeneAmp PCR system nomical conditions, considering that morphological char- Thermal Cycler (model 9700 Applied Biosystems, Fos- acteristics may he influenced by environment or cultural ter City, CA) in total 20-iL volume reactions following techniques (I-linrichsen et al. 2001). Each genotype was typical protocols (Dangl et at. 2005). Each sample was repeated five times. Accessions selected for study were analyzed at 17 SSR loci: VVMD5, VVMD7. VVMD2I, Cannaiola di Marta (CM VT!) and Cannaiola Macchie di VVMD24, VVMD25, VVMD27, VVMD28, VVMD3I, Marta (CMM VT2), collected in two vineyards of differ- VVMD32. VVMD34, VVMD 36 (Bowers et. al. 1996, ent age and cataloged as distinct accessions according to 1999), VrZAG62, VrZAG79. VrZAG83, VrZAG93 (Sefc the different local names and descriptions supplied by the et al. 1999), and VVS2 and VVS29 (Thomas and Scott donors Greco (G VT1), Greco verde (G VT2), Pagadebito 1993). Six internationally adopted reference markers were (PG VT5), and Rossetto (RS VT6), collected in various included (This et al. 2004). Forward primers were labeled parts of Tuscia Canaiolo nero (CN VT), present in the with one of three fluorescent dyes. Fragment amplifica- same collection and used as a test cultivar listed in the tions were verified on 2% agarose gels. Samples were pre- Italian Register of Grape Varieties. In addition to these pared for capillary electrophoresis by diluting 1.0 iL am- genotypes, four accessions of the same or similar name plified product and 0.4 1iL internal-size standard 400UD in the collection at the USDA National Clonal Germplasm ROX (Applied BiosystemS) in 12 iL formamide. Typically. Repository, Davis. California (NCGR-D) were sampled products from three loci labeled with different fluorescent for the analysis of DNA only (Canaiolo nero, Greco hi- dyes were multiplexed. Amplified fragments were sepa- anco, Pagadebito. and Moscato bianco). One accession rated by electrophoresis on an ABI Prism 3100 Genetic of Moscato (MS VTI) from DIPROV was also sampled Analyzer (Applied Biosystems) using 22 cm capillary with (Figure 1; Table 1). 3100 POP-4 as the matrix (Dangl et al. 2005). DNA extraction and microsatellite analysis. Young The microsatellite profiles obtained were compared leaves from near the shoot tip of rapidly growing grape- with previously reported profiles of vines in collections at vines were collected and rapidly dried between blotting NCGR-D and INRA (Domaine de Vassal. France) (Dangl paper in sealed, labeled envelopes, which were placed in plastic bags with —20 grams Drierite (W.A. Hammond et al. 2001). AmpelograPhical description. Ampelographic data Drierite, Xenia, OH). Total DNA was extracted from were collected during 2005 and 2006 as specified by the —20 mg dried leaf tissue using a DNeasy Plant Mini Kit Rossetto VT6 Greco VT1 Greco verde VT2 Pagadebito VT5 Cannaiola Marta VT1 Cannaiola M. Marta VT2 Leaf shapes of selected accessions used for ampelographic observations (obtained from real leaves). Figure 1 Am. J. Enol. Vitic. 60:1 (2009) 112— Muganu et al. Table 1 Accessions and cultivars used in the study. Organisation Internationale de la Vigne et du Viii (OIV Accession Code Sourcea Origin (Italy) 1983). Thirty-five morphological descriptors were used, including the preliminar y minimal descriptors relative to Cannaiola di Marta CM VT1 DIPROV Marta (Tuscia) shoots, young leaves, mature leaves, vine shoots, inflo- Cannaiola Macchie di Marta CMM VT2 DIPROV Marta (Tuscia) rescence, hunches, berries, and seeds. Ten readings per Canaiolo nero CN VT DIPROV Tuscany each descriptor were taken on five plants. Berry and seed Greco G VT1 DIPROV Vignanello (Tuscia) measurements were made at harvest using 50 berries, Greco verde G VT2 DIPRDV Vignanello (Tuscia) from 10 bunches, taken between 3/5 and 4/5 of hunch axis. Data oil main phenological phases (Baillod and Pagadebito PG VT5 DIPRDV Montefiascone (Tuscia) Baggiolini 1993) were collected during the two years. Rossetto AS VT6 DIPRDV Montefiascone (Tuscia) Moscato MS Vii DIPADV Montefiascone (Tuscia) Results and Discussion Canaiolo nero U6434 NCGR-D Ampelographical data. The Tuscian varieties selected Pagadebito F090a NCGR-D for this study were those most frequently mentioned in Greco bianco D044b NCGA-D historical sources. These sources also indicated that syn- Moscato
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