ABSTRACT

A renewed interest in inter-specific varieties has recently emerged, due mainly to producers and consumers more aware of organic farming and impact of phytochemicals in the environment. The assessment of 19 European Vitis hybrids was investigated in an area mostly dedicated to viticulture, the North-Eastern Italy. Major agronomic traits, yield, quality characteristics and disease resistance were evaluated during a three-year period (2004 to 2006). Wine sensory analyses were performed and compared with international Vitis vinifera varieties. Even though no genotypes resulted adequate for market release, the results obtained confirm the potential importance of hybrids in an “eco-friendly” viticulture and identify the genotypes interesting for further investigation and breeding: GF 138-3 and GA 48-12 showed good agronomic performance, resistance to more grape diseases and high quality wine.

1. INTRODUCTION

Successful viticulture must meet the requests of consumers and growers for good wine quality, disease and insect tolerance and low environmental impact [1,2]. Recurrent environmental issues have increasingly sparked political-social discussions over the last ten years. The European agricultural policies implemented guidelines focused on improved management strategies, integrated agronomic practices in the vineyard (2009/128/CE Directive) and a reduction in the use of pesticides and fungicides, using more disease-tolerant varieties in place of conventional ones. Hybrid varieties could be the most promising tool for low input, low cost and time-saving viticulture because of their tolerance to diseases and insects [3,4]. The wine industries in many extra-European Union countries currently use a high percentage of inter-specific varieties with good results and fund specific breeding programmes [5]. In Canada, USA, Switzerland, Germany and Hungary several inter-specific wines are commercialised.

Since 1960, inter-specific varieties had been used to successfully introgress tolerance to pests and diseases, such as powdery mildew (Erysphe necator Schwein), downy mildew (Plasmopara viticola) or phylloxera (Dactulosphaira vitifoliae Ficth.) [6-8]. These varieties are the result of efforts to combine the quality of traditional European varieties (Vitis vinifera) and pyramid different resistance traits typical of American varieties (Vitis riparia, Vitis labrusca, Vitis aestivalis, Vitis berlandieri and Vitis amurensis). Inter-specific breeding was especially important after the massive destruction of European vineyards, as consequence of the invasion of serious fungal diseases from the US during the second half of the 19^th Century. At the beginning of 20^th century, over 6000 hybrids were registered in Europe. Unfortunately, the offspring of these varieties often lose the stable yield and good quality traits of their European parents due to the complex polygenic base, which governs the resistance and the quality of the grapes [9,10], demonstrating that inter-specific breeding methods are quite unsuccessful. Finally, the diffusion of pesticides, the employment of the first rootstocks tolerant to phylloxera, the low quality of wines obtained and the possible presence of toxic metabolites have led to the unpopularity of hybrids [11,12]. Since then, crosses have been only performed in Germany [7,13], Austria [14], France [15] and Hungary [16]. In 1990, European wine area cultivated with hybrids was greatly reduced [0.04%, especially concentrated in Romania; 17]. Anyway, for many years, the Experimental Station for Viticulture (now CRA-VIT) in Conegliano (Treviso, Italy) investigated the quality of second-generation inter-specific hybrids [18,19]. In 1986, the first attempts to use in vitro plantlets to screen grape genotypes for resistance to disease infections were reported [20]. More recently, marker-assisted selection (MAS) has been use to greatly improve the introduction of genetic complex traits, such as grape quality and disease resistance, through reduced time and costs and avoiding the many problems that typically result from backcrosses [21-25].

Currently, the promising market of hybrids was first established through the cultivation of “Regent” in Germany, where it is grown over an area of more than 600 ha; a future similar trend in Europe, particularly in Italy (INFOAM 2000; Deutsches Weinbaujahrbuch 1995- 2003), can be foreseen. As soil, location and climate (typically referred together as Terroir) play a central role in vine performance and wine qualitative characteristics, the relationship between the hybrids and the environment is a crucial aspect that must be considered to evaluate their potential adoption in the investigated area. Thus, the aim of this study was to survey phenological, agronomic and qualitative performances, along with their resistance to the most common vine diseases, of 19 European wine grape hybrids.

2. MATERIALS AND METHODS

2.1. Site and Genotypes Description

The 19 inter-specific red and white wine grapes varieties, listed in Table 1 and present in the field collection of the CRA-VIT (Research Centre for Viticulture) in Conegliano (Treviso, Italy 45˚51'8.92''N, 12˚15'31.53''E)

were replanted in an experimental vineyard located near Motta di Livenza (Treviso, Italy 45˚47'5.91''N, 12˚35'38.90''E).

The hybrids were monitored for 3 years (2004 to 2006) and compared for agronomic parameters and disease resistance with “Pinot gris”, as this is an international cultivar widely cultivated in the area. The site is characterrised by a heavy soil (35% - 40% of clay). The climatic data accumulated for 3 years of study, are reported in Table 2. Approximately 11 - 13 chemical treatments per year against powdery and downy mildew are usually applied because the climate and soil conditions make this site habitat particularly suitable for high-severity fungal diseases. Anyway, to assess the resistance against fungal diseases, the varieties under investigation were treated only four times during the vegetative period, using formulations without copper before flowering and with copper hydroxide after flowering. Canopy and soil management were conducted according to practices commonly adopted in the vineyards of the area. The vine spacing was 3 × 1.7 m (1960 vines/ha), and the vines were trained using the Sylvoz system, with 3 canes of 10 - 12 buds each. The cultivars were grafted on the rootstock Kober 5 BB. The experimental design was replicated using three 15-plant blocks for each hybrid.

2.2. Analysis

The main phenological stages, such as budbreak, bloom and veraison (defined as 50% of plants upon the

Table 1. Inter-specific varieties investigated in the study: colour of berry, origin of varieties and their pedigree are reported.

Table 2. Average values of mean temperature (˚C) and rainfall (mm) in the three years under investigation.

stage) were evaluated and the variation quantified using standard deviation. Sugar accumulation and acid degradation were monitored upon the initiation of veraison (data not shown) to determine the ripening level and the harvest time. The shooting percentage (shoots/buds), potential fruitfulness (bunches/shoots) and real fruitfulness (bunches/buds) were determined for all the varieties. The real fruitfulness of the first four buds of each cane was also determined in 2005. All these parameters were compared among the hybrids and to “Pinot gris”. At harvest, the average cluster weight and yield per vine were determined. The grape quality was analysed by measuring the soluble solids, titratable acids and pH using a digital refractometer (ATAGO PR-101), a manual titrator (Digitrate Pro 500-Jencons) and a Crison pH metre, respectively.

Based on the agronomic and grape qualitative data, 0.3 tons of grapes from only the most promising varieties were harvested yearly and microvinified (Table 3). Single vinifications were performed with each treated grape. The winemaking process was identical for all vinification experiments. The assessment of the overall wine quality was performed using blind tasting.

Taste panelists evaluated the general organoleptic characteristics, such as aroma and flavour intensity, complexity, balance, structure and finesse.

The wines from two of the most widespread varieties cultivated in the area were used as standards for the sensory analysis. “Pinot blanc” was used as a standard for white wines, as it presents more neutral aroma than “Pinot gris”, and “Franconia” was used as a standard for the red wines. Both standard wines were obtained using

Table 3. List of varieties microvinified in the three years under investigation.

• marks up the year of microvinification.

grape from the experimental site and by using the same winemaking process adopted for the hybrids.

The damage caused by the most common diseases associated with North-Eastern Italy climate (Downy Mildew, Botrytis and Black Rot) was assessed at harvest in 2004 and 2005 using visual inspections and evaluating leaves and bunches separately. A total of 150 clusters and 200 leaves sampled from different parts of the grapevines for each replicate were used in the assessment. Infected leaves or bunches were ranked according to a scale: 0 = no symptoms; 1 = 1% - 10%; 2 = 10% - 25%; 3 = 25% - 50%; 4 = 50% - 75%; and 5 = 75% - 100% of infected leaf areas or infected berries per bunch, respectively. The data regarding disease severity were processed according to Townsend-Heuberger formula [26] to calculate the percentage of infection (I%):

where I% = percentage of infection; n = number of leaves or bunches in each class; v = class value; z = highest class value; N = total amount of assessed leaves or bunches. The data were statistically analysed using ANOVA and Duncan’s multiple range test, using the statistical package “Statistica 8.1”, (StatSoft Inc., 2007). The average, standard deviation and P were calculated.

3. RESULTS AND DISCUSSION

3.1. Phenological and Agronomic Description

The full phenological expressions of the hybrids demonstrated their adequate adaptation to the North Italian area. The mean dates for the three years, relating to the phenological stages, compared with “Pinot gris”, are reported in Figure 1. In the North-East Italian areas,

“Pinot gris” is considered as an early wine grape variety, registering budbreak in the first/second decade of April, flowering in the first 10 days of June, and veraison in the last decade of July. The phenological data showed a high variability among genotypes in the budbreak phase (Figure 1(a)), reflecting a higher and different sensitivity to spring weather compared with “Pinot gris”. The budding mean date of “SV 39639”, “GA 52-42”, “GF 64- 170-1”, “Ambror” and “Seibel 5178” occurred in the second decade of April. “Ambror” and “Seibel 5178” were the most stable varieties observed during budbreak. The earliest cultivars were “A × GM 64-94-5”, “Saphira”, “GA 48-12” and “Sirius”. On average, the blooming, veraison and harvest dates for these varieties all occurred later than observed with “Pinot gris”. At flowering, the delay was particularly evident for “Seyve-Villard 12390” and “GM 723-4”, which flowered 9 - 10 days after “Pinot gris”. Regarding the veraison date (Figure 1(b)), most varieties registered a 2-week delay compared with Pinot gris, occurring at the end of July and the beginning of August. Almost all hybrids exhibited a harvest time within the first two weeks of September (approximately 7 days after “Pinot gris”). “Staufer”, “Phoenix”, “Regent”, and “Seibel 7052” exhibited the earliest, completing the growing season at the end of August, together with “Pinot gris”, “Seyve-Villard 12390” was the only late variety, completing berry ripening approximately one month after “Pinot gris”. Thus, this hybrid was not suitable for the investigated area, as long phenological cycles might represent a limiting factor due to autumnal frosts and rainfall that could inhibit berry maturation. The hybrids showed a longer budbreak-veraison period (107 days compared with 97 for “Pinot gris”) and a quite short veraison-harvest phase (35 days compared with the 39 days observed for “Pinot gris”). The only exceptions were “Regent”, which exhibited late budding and precocious harvesting, and “Seibel 7052”, which was consistently precocious for all parameters tested. Shooting percentage, potential and real fruitfulness values are reported in Table 4. The shooting percentage was lower for the hybrids compared with “Pinot gris” (84.7% vs. 92.5%, respectively). In contrast, the values of fruitfulness were similar to “Pinot gris”. It was evident that no fruitfulness decrease was directly associated with the use of hybrids. However, “Seibel 7052”, “GM 7743-8” (Figure 2(a)) and “SV 39639” (Figure 2(b)) exhibited high fruitfulness potential, which frequently presented three bunches per shoot. Considering real and potential fruitfulness, “Seibel 7052”, “GM 7743-8” and “Seyve-Villard 39639” exhibited better performances than the V. vinifera cultivar, while “Seyve-Villard 12390” and “GA 52-42” exhibited the worst performance. Alaa Al-Joumayly [27] reported that fertility coefficients are genetically determined and only slightly conditioned according to season. Notably, the fruitfulness of the first basal buds is a relevant aspect to consider the manual and mechanical spur pruning [28, 29]. “Seibel 7052”, “Seyve-Villard 39639” and “GA 52- 42” were unsuitable for these methods of pruning because of their low fruitfulness in the first 4 buds (Table 4). The standard deviation showed a high variation among different years, particularly for budbreak percentage.

3.2. Grape Production and Quality

Cluster weight (g) and yield values (kg/vine) of these cultivars showed an opposite trend compared with “Pinot gris”, as cluster weight was, on average, lower (137 g) than that of “Pinot gris” (167 g; Table 5). “Seyve-Villard 12390” and “Villard Blanc” showed the highest cluster weight; however, “Seibel 5178”, “Phoenix”, “GF 138-3”, “GA 52-42” and “Seyve-Villard 39639” recorded the lowest values at nearly 100 g. The yield (tons per hectare)

Table 4. Budbrake (%), real fruitfulness, real fruitfulness of first 4 basal buds and potential fruitfulness of the 19 hybrids compared with “Pinot gris”. The data reported are average of the data registered in 2004, 2005 and 2006, and their variability was measured as standard deviations.

Note: Letters represents the significance of variability among the hybrids with p < 0.05 (Duncan test).

Table 5. Cluster weight and yield per vine of the 19 hybrids and “Pinot gris”. The data reported are average of the data registered in the three years, and their variability was measured as standard deviations.

Note: Letters represents the significance of variability among the hybrids with p < 0.05 (Duncan test).

was significantly high, with over 20 t/h obtained for the most productive hybrids and 4 - 5 t/ha obtained for the least productive varieties. On average, the hybrids produced more (8.1 kg/vine) than “Pinot gris” (7.3 kg/vine), suggesting fruitfulness, as reported by previous studies [30,31]. The most productive varieties were “Sirius”, “Regent”, “Seibel 7052”, “Orion”, “GA 48-12” and “GM 7743-8”, all presenting a yield greater 9 Kg/vine. Overall, we observed high variability among the different genotypes. Over the three-year period, the “Regent” and “Ambror” registered the highest variation. The soluble solids, titratable acids and pH values were measured to assess the grape quality [32]. A high variability was observed among the hybrids (Figure 3), which reflect differences in their genotypes and the annual climate conditions, consistent with the observations of Pavloušek and Kumšta [33], showing the strong effect of the year and cultivar on the pH values and titratable acid contents. “Ambror” showed the highest soluble solid values (20.1˚ Brix), followed by “Regent”, “Seibel 7052” and “GF 138- 3”, which all exhibited values superior to “Pinot gris”, “Seibel 5178” and “Saphira” were similar to “Pinot gris”

(18.1˚ Brix), while “Staufer” recorded the lowest value (14.8˚ Brix). A comparison of the yield per vine (Table 5) to the sugar richness (Figure 3) revealed that “Regent” and “Seibel 7052” had good production and high-soluble solids; no negative regression was observed among the hybrids between the two parameters. The estimated titratable acids values were primarily high for most hybrids (Figure 3(b)), which also reflected moderately low pH values (Figure 3(c)). “Seibel 7052”, “Phoenix”, “Sirius” and “GF 138-3” exhibited acids levels similar to “Pinot gris” (6 - 7 g/L). High acid values indicate a positive trait in the present global warming scenario and also represent desirable characteristics, particularly for sparkling wines. The increase of air temperature corre-

sponded with a similar increase in pH values in must. As Keller [34] suggested, the pH values of musts should not exceed 3.6 to avoid a decrease of quality of wine produced. As shown in Figure 3(c), all hybrids presented pH values below this limit.

Regarding the wine characteristics and quality, in 2004 no white wines obtained from these hybrids performed as well as the reference commercial variety (“Pinot blanc”; Figure 4(a)). The quality of “Phoenix”, “GA 52-42” and “Ambror” were among the best reported, mostly for the olfactory intensity with a pronounced aroma and an adequate complexity. These qualities indicated low finesse, which is a characteristic that was confirmed in the 2005 and 2006 tastings (Figures 4(c) and (e)). Overall, “GA 48-12” demonstrated the best performance (not included in the 2004 tasting), exhibiting scores similar to “Pinot blanc”. Among the remaining white varieties, “Ambror” achieved interesting results, particularly for olfactory intensity and taste persistence.

Concerning red wines obtained from hybrids, the quality was lower than the reference variety wine (“Franconia”) in all three years (Figures 4(b), (d) and (f)). In 2004 (Figure 4(b)), the black wines did not exhibit high parameters associated with the bouquet or taste, except for “GF138-3”, which showed appreciated values for the olfactory aromatic intensity. “Regent” exhibited increased values from the first to the third year. In 2006 (Figure 4(f)), the lowest yield (8.1 Kg/vine) and sensory characteristics for “Regent” were similar to “Franconia”, but the olfactory value was low.

3.3. Estimation of Disease Tolerance

The percentage of Downy Mildew, Botrytis and Black rot infection is reported in Table 6. During 2004 the rainfall and temperatures were standard for the area; hence, the antifungal treatments sufficiently limited disease damage. In 2005, high rainfall occurred during the blooming and berry-set periods, when the vines are more susceptible to Downey Mildew infections, and at the beginning of September, when more severe Botrytis infections occur. In the second year of the trial, “Saphira” and “Seibel 7052” were more susceptible to Downy Mildew than “Pinot gris”, while “SV 39639”, “SV 12390” and “Villard Blanc” were the most tolerant for leaf and cluster, exhibiting negligible symptoms of infection (Table 6). However, “Villard Blanc” showed the worst results for black rot, and “Phoenix” exhibited the highest percentage of infection for Botrytis (Table 6). Although each of the hybrids showed a high resistance to at least one of the diseases evaluated, it was not possible to identify a hybrid with a low percentage of infection for all three diseases. Even though “GF 138-3” showed a low disease incidence, “SV 12-390” resulted the most tolerant among the red wine cultivars. Whilst, three white wine cultivars, “GA 48-12”, “Ambror” and “Regent”, resulted adequately tolerant to Downy Mildew, consistent with the data reported in Germany [35] and in Poland [3].

4. CONCLUSIONS

Viticulture in North-Eastern Italy is difficult due to the high incidence of Downy Mildew and Botrytis, resulting from high rainfalls and air humidity. Unlike other European countries, e.g., France and Spain, Italy needs to drastically limit the spread of fungal diseases. The necessity of a more sustainable viticulture urgently requires the identification for new vine genotypes resistant to the more common vine diseases and the production of wine according to commercial qualitative demands. Specific

Table 6. Disease severity (expressed as % of infection according to Townsend-Heuberger formula) at harvest in 2004 and 2005.

Note: Letters represents the significance of variability among the varieties with p < 0.05 (Duncan test).

studies concerning the cultivation and potential comercial use of hybrids in North-Eastern Italy have not been previously conducted. Although stringent European Union rules strictly limit the use of these hybrids, and advantages and drawbacks of their employment are being discussed, the results obtained from the present study indicated that breeding programmes must be pursued.

None of the hybrids under study showed high values for all agronomical characteristics tested, nor a global quality sufficient to suggest their direct application in organic viticulture. Furthermore, the high variability among the genotypes observed for agronomic, phenological and qualitative performances could provide choices suitable to different and specific oenological goals. Among the red wine cultivars, “SV 12-390” showed the lowest infection, adequate production (due to greater weight of the bunch) and a good overall wine quality. However, basing on the results obtained in the present research, our opinion is that “GF 138-3” is the most promising hybrid, showing a strong olfactory intensity, an interesting overall sensory quality and a low disease incidence. Schwab [35] recommended “Regent” for organic viticulture in “Franconia” (Germany) because the sensory analysis rated this wine at a quality higher than Pinot noir. Indeed, “GF 138-3” shares a common pedigree with “Regent”, showing similar agronomic and qualitative characteristics. However, the results of the present study suggest that “GF 138-3” is better assessed in North Eastern Italy. Among the white wine varieties, our choice is “GA 48-12”, showing a high quality, even when it was not fully resistant to Downy mildew. Even though, the sensory analysis rated “GA 48-12” as interesting in tasting, as reported for other hybrids [36,37], its yield must be reduced to improve the sugar content.

The further improvement of wine quality before the commercial release of hybrids is necessary. Moreover, research on agronomic practices aimed at improving the performance of single genotypes could reinforce the results obtained through breeding programme.

5. ACKNOWLEDGEMENTS

Thanks are due to Dr Alessandro Zanzotto for his contribute in the disease evaluation.

REFERENCES