by Lisa Klima Johnson, Department of Horticulture, University of Georgia
Muscadines (Vitis rotundifolia Michx.) are a unique minor fruit crop native to the southeastern United States, related to the bunch grape, but with a musky flavor and several distinctive characteristics. They are eaten fresh, as well as processed into juice, wine, and preserves. Muscadines have become more popular recently due to an awareness of their high polyphenolics content. There are three species in the Muscadinia subgenus, and they are sometimes used for increasing genetic variation in the bunch grape subgenera, the Euvitis. Muscadines have 40 chromosomes (2n = 2x = 40), which differs from the 38 chromosomes in bunch grapes, and can cause graft incompatibility and difficulty in hybridization. Male muscadine vines are no longer used in commercial vineyards. Breeding programs focus on providing female or self-fertile improved varieties. Muscadines have several desirable qualities breeders try to incorporate: large berry size, self-fertile flowers, seedlessness, good flavor, dry picking scar, early maturity, thin or palatable skin, high soluble solids, firmness, eye appeal, vigorous growth, and resistant to disease. Traditional breeding is being used to develop new cultivars, but some molecular breeding techniques are proving to be useful, especially involving disease resistance. Muscadine vines are evaluated over a period of several years, and selection can be complicated due to the large range of characteristics desired in a new cultivar. The primary challenge in breeding muscadines is that disease resistance and vigor decrease as the fruit quality increases, which provides an excellent subject for a lifetime of fruit breeding work.
Muscadines (Vitis rotundifolia Michx.) have a distinct flavor from bunch grapes, a thick skin, and large seeds. They are considered to be more fruity and musky (Morris and Brady, 2004). Most often, muscadines are eaten fresh, or used for producing wine and preserves. Wine and juice products have become more popular in recent years, and the fresh fruit has always been a regional favorite of the southeast (Conner, 2009). Also, interest has grown in using dried muscadine pomace as a functional food or nutritional supplement, due to the high levels of polyphenolics in the skins and seeds (Morris and Brady, 2004; Vasanthaiah et al., 2011; Vashisth et al., 2011). Pomace is the skins, seeds, and pulp left over from wine or juice processing. The breeding of muscadines is complex since it requires selection of several important traits at once, and a significant amount of time is required to develop new cultivars. Both molecular and traditional breeding techniques are being used in the advancement of this popular minor fruit crop (Conner, personal communication, 2012).
Species, Origin, and Genetics
Muscadines are a member of the genus Vitis, which is divided into two subgenera: Euvitis and Muscadinia. The area between the Black Sea and Caspian Sea is thought to be the center of origin of Vitis (Basiouny and Himelrick, 2001). Euvitis consists of the more common bunch grapes, Vitis vinifera, as well as several other species. Muscadinia consists of three species which are native to the southeastern part of the United States, and parts of South America. Vitis rotundifolia Michx. (muscadine) is the species which is grown commercially and found commonly in the wild, and will be discussed here. The two others, Vitis munsoniana and Vitis popenoeii, are found in very limited ranges, and are used occasionally to increase variation, but are not commercially important species (Conner, personal communication, 2012). Muscadines have been observed in North America since the 1500s in North Carolina (Ison, 1988; Morris and Brady, 2004). Wild and cultivated muscadines are found in southeastern states north to Virginia, Tennessee, and Arkansas, and west to east Texas (Basiouny and Himelrick, 2001). The first self-fertile muscadine was noted in 1910, also in North Carolina (Ison, 1988). Muscadines are a familiar crop in this region, but not nationally. The 'Scuppernong' is commonly considered to be a different fruit than a muscadine, but it is simply a cultivar. Many bronze-colored cultivars are usually called 'Scuppernong', as it is the most famous and oldest cultivar with that characteristic. However, there have been several bronze releases like 'Tara' (Lane, 1993) and 'Florida Fry' (Mortensen et al., 1994a) (Figure 1). The Muscadinia subgenera has 40 chromosomes (2n = 2x = 40), in contrast to the Euvitis subgenera, which has 38. This causes difficulty in hybridization between the two groups (Janick and Moore, 1975), and they are graft-incompatible as well (Olien, 1990). Often, the Muscadinia is viewed as a source for potential genetic variation for the Euvitis group in spite of these difficulties (Janick and Moore, 1996). This family is tolerant to a wide variety of diseases, is vigorous, and adapted to hot climates. Seven V. rotundifolia accessions, as well as three hybrids with V. rotundifolia as one of the parents, were found to be resistant to downy mildew, a commercially important grape disease (Staudt and Kassemeyer, 1995). Three V. rotundifolia accessions, as well as seven hybrids with V. rotundifolia as one of the parents, were found to be resistant to powdery mildew, another serious grape disease (Staudt, 1997). Disease resistance from muscadines is a coveted trait to grape breeders, but the flavor profile does not match Euvitis standards for fresh market fruit or wine production (Riaz et al., 2009).
In the wild, muscadines are most often found to be dioecious, with 50 - 75% of plants being male (Ison, 1988). In commercial vineyards, female cultivars are usually interspersed with a perfect-flowered pollinizer. Male pollinizers will not bear fruit, making the perfect-flowered types preferable. Male plants are not used in modern production. Most of the fruit set on female cultivars, and close to half of the fruit set on self-fertile cultivars is attributable to cross-pollination, which is facilitated by both insects and wind (Sampson et al., 2001). Muscadine flowers occur in racemose panicles, with 10 - 30 small, green, hypogynous flowers (Basiouny and Himelrick, 2001; Rieger, 2006; Westwood, 1993). A calyptra, or cap, remains attached to the flower until anthesis, and may stay on afterwards in some cases, promoting self-pollination over cross pollination in self-fertile cultivars and preventing pollination in female cultivars (Conner, personal communication, 2012) (Figure 2). Most of the pistils become receptive only after the calyptra abscises (Sampson et al., 2001). Some cultivars have pistillate flowers, in which the anthers are short and reflexed, and the perfect-flowered varieties have anthers that extend past the pistil (Conner, personal communication, 2012; Rieger, 2006) (Figure 3, Figure 4). The inheritance of sex in grapes is somewhat controversial, but most research indicates that sex is determined by a single locus with three alleles with a decreasing order of dominance: M (male flowers), H (hermaphroditic flowers), and F (female flowers) (Reisch and Pratt, 1996).
Basiouny and Himelrick (2001) describe 119 muscadine cultivars, 64 of which no longer hold commercial value. The University of Georgia muscadine breeding program has released over 30 cultivars since 1909 (Conner, 2010). One of the most popular commercial cultivars was a UGA release in 1971, 'Fry', which is a female, bronze-fruited variety (Basiouny and Himelrick, 2001; Conner, 2010). The top 12 cultivars in production in the southeast are 'Carlos', 'Dixie', 'Doreen', 'Fry', 'Jumbo', 'Magnolia', 'Nesbitt', 'Noble', 'Regale', 'Summit', 'Triumph', and 'Welder' (Basiouny and Himelrick, 2001). A Euvitis x V. rotundifolia hybrid, 'Southern Home', was released in 1994. It was promoted primarily for home use, and its ornamental value, vine vigor, and disease resistance were highlighted (Mortensen et al., 1994b) (Figure 5). This hybrid features mainly muscadine qualities, with a small berry size, but the flavor is not suitable for commercial production (Conner, 2010). The deeply lobed leaves typical of V. vinifera are visible on 'Southern Home' (Figure 5). This leaf type is often used as a visual marker to indicate a successful hybrid cross with V. vinifera (Conner, personal communication, 2012). New cultivars are periodically released, like 'Southern Jewel', and 'Delicious', however, each new cultivar has a group of characteristics that still does not match all the qualities desired by growers (Gray et al., 2009a; 2009b).
Fruit breeding is not successful unless the desired combination of traits is available in one cultivar. It is critical to select for several traits simultaneously. Muscadine breeding is not about trying to add one trait to an already otherwise perfect cultivar. No cultivars available in muscadine have all the qualities growers want (Basiouny and Himelrick, 2001). The most desirable qualities for growers are large berry size, self-fertile plants, seedlessness, and great flavor. Many qualities also need to be present for a cultivar to be popular which may not be obvious. For example, berries should have a dry picking scar. This refers to the area of the abscission zone on the fruit, where the pedicel was attached prior to harvest. The fruit should abscise cleanly from the pedicel, leaving a healed, dry picking scar, so the flesh is not exposed and available for bacterial infection postharvest. Berry clusters should be large and should not shatter. Vines need to be vigorous and resistant to disease. Berries should mature early, have thin or at least palatable skin, display high soluble solids and firmness, hold up well during storage, and especially have good eye appeal (Conner, personal communication, 2012; Morris and Brady, 2004; Stringer et al., 2008).
Cultivar selection will be made easier for the grower over time with the introduction of more perfect-flowered varieties; however, the fruit quality is always of utmost importance. In fresh fruit production, large fruit size is desirable, as well as early maturity, synchronous harvest time, non-shattering berries, and dry picking scar (Conner, personal communication, 2012; Vasanthaiah et al., 2011). For pick-your-own operations, similar qualities are important, but a range of harvest times stretches the harvest out over a longer period of time. In wine or juice processing vineyards, synchronous harvest time is important so all the fruit can be harvested and processed efficiently. The muscadine juice color is also critical, as an undesirable change can occur during processing (Morris and Brady, 2004; Vasanthaiah et al., 2011). Fruit size is not as critical. In all types of operations, disease resistance is of utmost importance. Seedlessness has long been a desirable quality, but it has not yet been possible (Olien, 1990).
In Georgia, traditional breeding is commonly used to improve muscadines. Two cultivars are selected for crossing which have the promise of a good combination of qualities. Since male vines are undesirable, the crosses are generally made between female and self-fertile varieties which normally gives a 1:1 self-fertile:female ratio in the resulting seedling progeny (Conner, personal communication, 2012) (Figure 3, Figure 4, and Figure 6). Self-fertile is one of the most desirable qualities in muscadines, but if only self-fertile vines are available, emasculation becomes a critical technique in breeding. Emasculation is difficult in muscadine due to the fragility of the clusters and the small size of the stamens and anthers (Conner, personal communication, 2012). For this reason, female vines are selected from progenies for use as parents while self-fertile vines are selected for use as parents as well as potential new cultivars. Flower clusters are bagged prior to anthesis (Figure 7). Perfect flowers are used whole, brushing their anthers on the stigma of the female flowers. Alternately, pollen can be collected and brushed onto the stigma surface, which is a technique that is used when the two varieties have different bloom times (Conner, personal communication, 2012). Clusters are re-bagged after pollination. Once fruit is mature, the seeds are collected and stratified for a period of three to six months. The seeds are planted in the greenhouse in the early spring and seedlings are transferred to the field in early- to mid-summer (Conner, personal communication, 2012) (Figure 8).
Phenotyping, Data Collection, and Selection
A hundred or more seedlings from a specific cross are grown on trellises similar to maturing vines, but with close spacing of about two feet (Figure 9). A large planting hole with rough sides has been shown to be important to muscadine development (Olien, et al., 1993). In two to three years, these vines will begin fruiting, and they will begin to be evaluated annually. If the cross appears to have produced a population which will produce several selections, the seedlings are evaluated for up to four years, and each seedling is evaluated weekly. Phenotyping reduces the seedling population quickly by ruling out plants without the critical qualities of fruit size, vigor, disease resistance, thin skin, and good flavor. Selections are made based on these first few qualities, and often the vine on either side of the selection is removed to give more space to the selected vine. At this point, selections are numbered and propagated, and a 2-vine plot is used to collect qualitative data (Figure 10, Figure 11). Estimates are noted of fruit size, scar qualities, skin type, maturity date, flavor and sweetness, shattering, cluster size, and disease resistance. As these selections are evaluated over a few years, the population with desirable qualities continues to shrink. Once the choice selections with cultivar potential are determined out of this group, they go into a replicated trial at three locations, one being a commercial vineyard, and the other two being UGA vineyards. Quantitative data is recorded on yield, harvest date, fruit size, soluble solids content, vigor, berry firmness, berry rot, and storage capability (Figure 12). Varying management practices at the three locations contribute to minor differences in final yield and quality data. Growers are invited to field days where they provide their opinion of the selections (Conner, personal communication, 2012).
Limited biotechnology techniques are in use with muscadines, probably owing to their status as a minor crop and the fact that the genome is not yet sequenced. Slightly over 50 nucleotide sequences and 12 expressed sequence tags (ESTs) have been stored with the National Center for Biotechnology Information database, and a decision has been made to use 'Noble' for genome sequencing and EST generation (NCBI, 2012; Vasanthaiah et al., 2011). Randomly amplified polymorphic DNA analysis confirmed that muscadines are significantly different genetically from bunch grapes, and that there is very little genetic variation among muscadine varieties (Qu et al., 1996). Also, the single sequence repeat marker profile of four muscadine cultivars indicated a difference from their published pedigrees, which probably means a mistake was made in identification (Riaz et al., 2008).
Muscadine is seen as a potential source for disease resistance for V. vinifera. One gene that has drawn some attention is the Resistance to Uncinula necator 1 gene (Run1), which is the grapevine powdery mildew resistance gene. Three molecular markers were confirmed as good candidates for Run1 marker assisted selection (Pauquet et al., 2001). Barker et al. (2005) found in their study that reduced recombination in the area of Run1 will require large populations in order to do fine mapping of this locus. Eibach et al. (2007) recommend selection using phenotypic data and molecular markers to identify resistance to powdery mildew and downy mildew. Gene pyramiding was used to combine Run1 with Ren1, which is another powdery mildew resistant gene found in a V. vinifera cultivar, and it was confirmed in a V. rotundifolia x V. vinifera BC5 hybrid family (Katula-Debreceni et al., 2010). The expression of several anthracnose-related defense genes was analyzed in 40 cultivars, in order to confirm phenotypic data (Louime et al., 2011).
Releasing a Cultivar
Once a selection has been thoroughly evaluated and is ready for release, a row is established so that nurseries can be provided with cuttings. The rooting percentage is also established and provided to nurseries (Conner, personal communication, 2012) (Figure 13). The cultivar should be available in limited quantities for a few years while nurseries build up their stock plants, and will be fully available in 3-4 years. Often growers discover problems with new cultivars that weren’t evident in initial testing, and probably only one out of five cultivars will be popular enough to be planted repeatedly after an initial test by growers (Conner, personal communication, 2012).
A few challenges slow the success of muscadine breeding in Georgia. The most common challenge is revealed as a trend: as the fruit quality increases, the disease resistance and vine vigor decreases (Conner, personal communication, 2012) (Figure 14). For example, when the skin of the fruit becomes thinner and more palatable, the fruit is more susceptible to fruit rot, or attack by beetles. The correlation between grape berry skin qualities and resistance to Botrytis cinerea was investigated because a tough skin is associated with resistance (Gabler et al., 2003). There was one V. vinifera x V. rotundifolia hybrid in the study classified as being highly resistant, displaying a high amount of cuticle, correlating to B. cinerea resistance (Gabler et al., 2003). Interestingly, it also had the highest number of lenticels in the study, which was not correlated with resistance, but may contribute otherwise to the difference between grapes and muscadines (Gabler et al., 2003). The other major challenge comes from the number one trait growers want, self-fertile flowers. Often, self-fertile cultivars set more fruit than the vine can support. This can result in a low soluble solids accumulation, small size, late maturity, and other undesirable features (Conner, personal communication, 2012). Heavier pruning or fruit thinning techniques may alleviate this problem and need to be researched on these cultivars. One positive trait coming out of a cross cancels out one or more other positive traits. Muscadine breeding continues to hold all the same challenges as breeding of other fruit crops, such as a long juvenile period and one generation per year. The advantage of muscadine breeding, as with some other fruit, is that there is no need to create inbred lines (Conner, personal communication, 2012). Muscadines are vegetatively propagated so once a desirable vine is discovered it is a trivial matter to produce a large number of clones. Phenotyping can be a lot of work, as in breeding any crop, but at least with muscadines, it can be a pretty sweet job (Conner, personal communication, 2012) (Figure 15).
Barker, C.L., T. Donald, J. Pauquet, M.B. Ratnaparkhe, A. Bouquet, A.F. Adam-Blondon, M.R. Thomas, and I. Dry. 2005. Genetic and physical mapping of the grapevine powdery mildew resistance gene, Run1, using a bacterial artificial chromosome library. Theor. Appl. Genet. 111: 370-377.
Basiouny, F.M., and D. G. Himelrick, editors. 2001. Muscadine Grapes. ASHS Press, Alexandria, VA.
Conner, P.J. 2009. Performance of muscadine grape cultivars in southern Georgia. Journal of the American Pomological Society. 63:101-107.
Conner, P.J. 2010. A century of muscadine grape (Vitis rotundifolia Michx.) breeding at the University of Georgia. Journal of the American Pomological Society. 64: 78-82.
Eibach, R., E. Zyprian, L. Welter and R. Topfer. 2007. The use of molecular markers for pyramiding resistance genes in grapevine breeding. Vitis. 46:120-124.
Gabler, F.M., J.L. Smilanick, M. Mansour, D.W. Ramming and B.E. Mackey. 2003. Correlations of morphological, anatomical, and chemical features of grape berries with resistance to Botrytis cinerea. Phytopathology. 93:1263-1273.
Gray, D.J., Z.J.T. Li, S.A. Dhekney, D.L. Hopkins and C.A. Sims. 2009a. 'Delicious': An early-ripening, self-fertile, multipurpose black-fruited muscadine grape. HortScience. 44:200-201.
Gray, D.J., Z.T. Li, S.A. Dhekney, D.L. Hopkins and C.A. Sims. 2009b. 'Southern Jewell': A self-fertile, black muscadine grape with fruit produced on bunches. HortScience. 44:1476-1477.
Ison, W.G. 1988. All About Muscadines Blueberries Blackberries for the Growers of these Fine Berries. Copyright 1988 by William G. (Bill) Ison.
Janick, J., and J. N. Moore, editors. 1975. Advances in Fruit Breeding. Purdue University Press, West LaFayette, IN.
Janick, J., and J. N. Moore, editors. 1996. Fruit Breeding. John Wiley & Sons, Inc., New York, NY.
Katula-Debreceni, D., A.K. Lencses, A. Szoke, A. Veres, S. Hoffmann, P. Kozma, L.G. Kovacs, L. Heszky, and E. Kiss. 2010. Marker-assisted selection for two dominant powdery mildew resistance genes introgressed into a hybrid grape population. Sci. Hortic. 126:448-453.
Lane, R.P. 1993. Tara Muscadine Grape. HortScience 28:232-232.
Louime, C., J. Lu, O. Onokpise, H.K.N. Vasanthaiah, D. Kambiranda, S.M. Basha, and H.K.Yun. 2011. Resistance to Elsinoe ampelina and expression of related resistant genes in Vitis rotundifolia Michx. grapes. Int. J. Mol. Sci. 12:3473-3488.
Morris, J.R., and P.L. Brady. 2004. The muscadine experience: adding value to enhance profits. AAES Research Report 982. Arkansas Agricultural Experiment Station. Fayetteville, Arkansas.
Mortensen, J.A., J.W. Harris, and D.L. Hopkins. 1994a. 'Florida Fry': a bronze muscadine grape. HortScience 29:1373-1374.
Mortensen, J.A., J.W. Harris, D.L. Hopkins, and P.C. Andersen. 1994b. `Southern Home': an interspecific hybrid grape with ornamental value. HortScience 29:1371-1372.
National Center for Biotechnology Information (NCBI). 2012. Nucleotide Database. http://www.ncbi.nlm.nih.gov/nuccore (accessed 16 Oct. 2012).
Olien, W.C. 1990. The muscadine grape: botany, viticulture, history, and current industry. HortScience 25:732-739.
Olien, W.C., C.P. Hegwood, and J.M. Spiers. 1993. Planting methods affect early growth and root distribution of muscadine vines. HortScience 28:1089-1091.
Pauquet, J., A. Bouquet, P. This and A.F. Adam-Blondon. 2001. Establishment of a local map of AFLP markers around the powdery mildew resistance gene Run1 in grapevine and assessment of their usefulness for marker assisted selection. Theor. Appl. Genet. 103:1201-1210.
Qu, X.P., J. Lu, and O. Lamikanra. 1996. Genetic diversity in muscadine and American bunch grapes based on randomly amplified polymorphic DNA (RAPD) analysis. J. Am. Soc. Hortic. Sci. 121:1020-1023.
Reisch, B. and C. Pratt. 1996. Grapes, p. 297-369. In:J. Janick and J.N. Moore (eds.). Fruit Breeding, Volume II: Vine and Small Fruit Crops. John Wiley & Sons, Inc., New York.
Riaz, S., A.C. Tenscher, R. Graziani, A.F. Krivanek, D.W. Ramming, and M.A. Walker. 2009. Using marker-assisted selection to breed Pierce's disease-resistant grapes. Am. J. Enol. Vitic. 60:199-207.
Riaz, S., A.C. Tenscher, B.P. Smith, D.A. Ng, and M.A. Walker. 2008. Use of SSR markers to assess identity, pedigree, and diversity of cultivated muscadine grapes. J. Am. Soc. Hortic. Sci. 133:559-568.
Rieger, M. 2006. Introduction to fruit crops. Haworth Food & Agricultural Products Press, Binghamton, NY.
Sampson, B., S. Noffsinger, C. Gupton, and J. Magee. 2001. Pollination biology of the muscadine grape. HortScience 36:120-124.
Staudt, G. 1997. Evaluation of resistance to grapevine powdery mildew (Uncinula necator [Schw.] Burr., anamorph Oidium tuckeri Berk.) in accessions of Vitis species. Vitis 36:151-154.
Staudt, G. and H.H. Kassemeyer. 1995. Evaluation of downy mildew resistance in various accessions of wild Vitis species. Vitis 34: 225-228.
Stringer, S.J., D.A. Marshall, B.J. Sampson, and J.M. Spiers. 2008. Performance of muscadine grape cultivars in southern Mississippi. HortTechnology 18:726-733.
Vasanthaiah, H.K.N., D. Thangadurai, S.M. Basha, D.P. Biradar, D. Kambiranda, and C. Louime. 2011. Muscadiniana, In: Kole, C., editor. Wild Crop Relatives: Genomic and Breeding Resources : Temperate Fruits. Springer-Verlag, Berlin. 2011. p. 65.
Vashisth, T., R.K. Singh, and R.B. Pegg. 2011. Effects of drying on the phenolics content and antioxidant activity of muscadine pomace. LWT--Food Sci. Technol. 44:1649-1657.
Westwood, M.N. 1993. Temperate-Zone Pomology. 3 ed. Timber Press, Portland, OR.