Vitis Riparia Research Articles

PHYLOGENETICS ANALYSIS OF NORTH AMERICAN NATIVE 'CYNTHIANA'/'NORTON' GRAPE CULTIVAR USING DNA MICROSATELLITE MARKERS

‘Cynthiana’/‘Norton’ is considered the best American native grape variety for fine wine with suspected to have tolerance to PD (Pierce’s disease) and proven low susceptibility to foliar and fruit disease. The variety has been very successfully grown for commercial wine production for more then a century in the eastern United States and for the past decade in Louisiana within PD zone as well. Most of the grapevine varieties in existence today are centuries old and are considered to have arisen by various means: domestication of wild vines, spontaneous crosses between wild vines and varieties, and crosses between two or more varieties. Using the methods of molecular analysis, the pedigree describing the genetic history of grape variety can be reconstructed. DNA microsatellites have proved to be the markers of choice for this purpose since they are transmitted in a codominant Mendelian manner. It is assumed that ‘Cynthiana’/‘Norton’ is originated from Vitis aestivalis, Michaux. We are investigating the phylogenetics of the ‘Cynthiana’/‘Norton’ grape via data mining in the North American grape germplasm collections, ampelographic analysis and specifically expressed in the variety microsatellite markers. We are using an already-published set of ten pairs of SSR primers designed for Vitis riparia. For the purpose developing more specific markers, ‘Norton’ publicly available expressed sequence tags (ESTs) were used to screen for SSR. Two 202 SSRs out of 2,101 ESTs were located and the promising SSR primers were synthesized for specific amplification in the targeted grape accessions. This study is designed to identify the pedigree and genetic make up of ‘Cynthiana’/‘Norton’ grape and to verify that this particular grape variety carry tolerance to Pierce’s disease and is appropriate for high quality red wine production under Florida environmental conditions.

Grapevine xylem sap enhances biofilm development byXylella fastidiosa

Xylella fastidiosa is able to form biofilms within xylem vessels of many economically important crops. Vessel blockage is believed to be a major contributor to disease development caused by this bacterium. This report shows that Vitis riparia xylem sap increases growth rate and induces a characteristic biofilm architecture as compared with biofilms formed in PD2 and PW media. In addition, stable cultures could be maintained, frozen and reestablished in xylem sap. These findings are important as xylem sap provides a natural medium that facilitates the identification of virulence determinants of Pierce's disease.

Influence of grapevine leafroll associated viruses (GLRaV-2 and -3) on the fruit composition of Oregon Vitis vinifera L. cv. Pinot noir: Free amino acids, sugars, and organic acids

Individual free amino acids, yeast assimilable amino acid (YAN) content, ammonia, organic acids, and simple sugars of berries from vines infected with GLRaV-2 or -3 were compared with paired vines free of these viruses. Samples were taken from two commercial vineyards during two growing seasons (2005 and 2006), with three different rootstock/scion combinations. Vines infected with GLRaV-2 did not differ significantly from their healthy counterparts in individual free amino acids, ammonia, or YAN content. Vines infected with GLRaV-3 were significantly lower in valine and methionine from Vitis riparia rootstock/‘Pinot noir’ clone 114 (VY2a) samples, and lower in glutamic acid from self-rooted/‘Pinot noir’ clone Pommard (VY2b) samples, compared to samples from their healthy counterparts. Samples from VY2b (self-rooted/‘Pinot noir’ clone Pommard) infected vines had significantly lower levels of malic acid and total organic acids compared to samples from their healthy counterparts. There were no significant differences between healthy and infected vines from all three rootstock/scion pairs in ammonia or free amino acids in samples taken during the weeks before ripening and at commercial harvest. This is the first study to report the influence of GLRaV-2 and -3 on ‘Pinot noir’ berries nitrogen (N) compounds significant to fermentation. Individual free amino acids may be inferior to phenolic compounds as indicators of GLRaV infection status. Published by Elsevier Ltd.

Mapping of Photoperiod-induced Growth Cessation in the Wild Grape Vitis riparia

In grapevines (Vitis spp.), the timing of growth cessation in the fall is an important aspect of adaptation and a key objective in breeding new grape cultivars suitable for continental climates. Growth cessation is a complex biological process that is initiated by environmental cues such as daylength and temperature, as well as water and nutrient availability. The genetic control of growth cessation in grapevines was studied by mapping quantitative trait loci (QTL) in a hybrid grape population. An F2 mapping population was developed by selfing a single F1 plant derived from a cross between an accession of the North American species Vitis riparia and the Vitis hybrid wine cultivar Seyval (Seyve-Villard 5–276). A linkage map was constructed using 115 simple sequence repeat (SSR) markers and six candidate genes in a population of 119 F2 progeny. The markers provided coverage of the 19 Vitis linkage groups with an average distance between markers of 8.4 cM. The critical photoperiod for growth cessation in lateral buds for the parents and F2 progeny was determined in a replicated field trial in 2001 and 2002 and under controlled photoperiod treatments in a greenhouse in 2002, 2003, and 2004. QTL analysis using composite interval mapping identified a single major QTL in the field and greenhouse trials. However, the field and greenhouse QTL mapped to different linkage groups in the two different environments, suggesting the presence of additional, nonphotoperiodic cues for induction of growth cessation in the field. In the greenhouse, where noninducing temperatures were maintained, a QTL on linkage group (LG) 13 explained 80.0% to 96.6% of the phenotypic variance of critical photoperiod for growth cessation. In the field, where vines experienced natural fluctuations in temperature and rainfall in addition to the naturally decreasing photoperiod, a QTL on LG 11 explained 85.4% to 94.3% of phenotypic variance.

Genetic dissection of sex determinism, inflorescence morphology and downy mildew resistance in grapevine

A genetic linkage map of grapevine was constructed using a pseudo-testcross strategy based upon 138 individuals derived from a cross of Vitis vinifera Cabernet Sauvignon x Vitis riparia Gloire de Montpellier. A total of 212 DNA markers including 199 single sequence repeats (SSRs), 11 single strand conformation polymorphisms (SSCPs) and two morphological markers were mapped onto 19 linkage groups (LG) which covered 1,249 cM with an average of 6.7 cM between markers. The position of SSR loci in the maps presented here is consistent with the genome sequence. Quantitative traits loci (QTLs) for several traits of inflorescence and flower morphology, and downy mildew resistance were investigated. Two novel QTLs for downy mildew resistance were mapped on linkage groups 9 and 12, they explain 26.0-34.4 and 28.9-31.5% of total variance, respectively. QTLs for inflorescence morphology with a large effect (14-70% of total variance explained) were detected close to the Sex locus on LG 2. The gene of the enzyme 1-aminocyclopropane-1-carboxylic acid synthase, involved in melon male organ development and located in the confidence interval of all QTLs detected on the LG 2, could be considered as a putative candidate gene for the control of sexual traits in grapevine. Co-localisations were found between four QTLs, detected on linkage groups 1, 14, 17 and 18, and the position of the floral organ development genes GIBBERELLIN INSENSITIVE1, FRUITFULL, LEAFY and AGAMOUS. Our results demonstrate that the sex determinism locus also determines both flower and inflorescence morphological traits.

Variation in recombination rates across Vitis species

Recombination rate data are presented for three populations of grape based on framework genetic linkage maps developed with simple-sequence repeat markers. These linkage maps were constructed from different Vitis species and represent three genetic backgrounds. The first population is pure Vitis vinifera, derived from a cross of the European cultivars Riesling and Cabernet Sauvignon. The second is an interspecific cross between two commercially used rootstock cultivars of different North American Vitis species parentage, Ramsey (Vitis champinii) and Riparia Gloire (Vitis riparia). The third population, D8909-15 (Vitis rupestris × (Vitis arizonica/Vitis girdiana form)) × F8909-17 (V. rupestris × (V. arizonica/Vitis candicans form)), is an F1 from two half-sibs. Genome-wide and chromosome-wide recombination rates varied across the three populations and among the six Vitis parents. Global recombination rates in the parents of the third F1 population, with a complex Vitis background, were significantly reduced. In the first and third populations, the recombination rate was significantly greater in the male parent. Specific genome locations with frequent heterogeneity in recombination were identified, suggesting that recombination rates are not equal across the Vitis genome. The identification of regions with suppressed or high recombination will aid grape breeders and geneticists who rely on recombination events to introgress disease resistance genes from the genomes of wild Vitis species, develop fine-scale genetic maps, and clone disease resistance genes.

Pathogen‐tested material of grapevine varieties and rootstocks

Pathogen‐tested material of grapevine varieties and rootstocks

Identification and Field Evaluation of Grape Shoot Volatiles Attractive to Female Grape Berry Moth (Paralobesia viteana)

Solid-phase microextraction (SPME) and gas chromatography coupled with electroantennographic detection (GC-EAD) were used to identify volatile compounds from shoots of riverbank grape (Vitis riparia) that attract the female grape berry moth (GBM, Paralobesia viteana). Consistent EAD activity was obtained for 11 chemicals: (Z)-3-hexen-1-yl acetate, (E)-linalool oxide, (Z)-linalool oxide, nonanal, linalool, (E)-4,8-dimethyl-1,3,7-nonatriene, methyl salicylate, decanal, beta-caryophyllene, germacrene-D, and alpha-farnesene. In flight-tunnel tests that involved female GBM and rubber septa loaded with subsets of these 11 compounds, we found that both the 11-component blend and a seven-component blend, composed of (E)-linalool oxide, (Z)-linalool oxide, nonanal, (E)-4,8-dimethyl-1,3,7-nonatriene, decanal, beta-caryophyllene and germacrene-D, elicited equivalent levels of upwind flight as freshly cut grape shoots. The removal of any of the seven compounds from the seven-component blend resulted in a significant decrease in female upwind flight responses. In a field trial with these two synthetic blends, traps equipped with either blend captured more female GBM compared to traps baited with hexane only (control), although the number of females caught was generally low. There were no differences in the number of males captured among treatments. Although in flight-tunnel trials, moths readily flew upwind to both grape shoots and rubber septa loaded with the best lures, they landed on shoots but not on rubber septa. Coupled with relatively low field catches, this suggests that additional host finding cues need to be identified to improve trap efficacy.

Transcript profiling in Vitis riparia during chilling requirement fulfillment reveals coordination of gene expression patterns with optimized bud break

Endodormant grapevine buds require a period of chilling before they break and begin to grow. Custom Vitis bud cDNA microarrays (9,216 features) were used to examine gene expression patterns in overwintering Vitis riparia buds during 2,000 h of 4 degrees C chilling. Three-node cuttings collected concurrently with buds were monitored to determine dormancy status. Chilling requirement was fulfilled after 1,500 h of chilling; however, 2,000 h of chilling significantly increased the rate of bud break. Microarray analysis identified 1,469 significantly differentially expressed (p value < 0.05) array features when 1,000, 1,500, and 2,000 h of chilling were compared to 500 h of chilling. Functional classification revealed that the majority of genes were involved in metabolism, cell defense/stress response, and genetic information processing. The number of significantly differentially expressed genes increased with chilling hour accumulation. The expression of a group of 130 genes constantly decreased during the chilling period. Up-regulated genes were not detected until the later stages of chilling accumulation. Hierarchical clustering of non-redundant expressed sequence tags revealed inhibition of genes involved in carbohydrate and energy metabolism and activation of genes involved in signaling and cell growth. Clusters with expression patterns associated with increased chilling and bud break were identified, indicating several candidate genes that may serve as indicators of bud chilling requirement fulfillment.

Influence of grapevine leafroll associated viruses (GLRaV-2 and -3) on the fruit composition of Oregon Vitis vinifera L. cv. Pinot noir: Phenolics

Some of the 10 known grapevine leafroll associated viruses (GLRaVs) have negative impacts upon vine productivity and grape quality, though these negative influences are dependent on factors such as GLRaV strain, cultivar, clone, rootstock, and vine age. This is the first study to report on GLRaV-2 and GLRaV-3 infected vines, with regard to phenolic compounds, and other fruit maturity indices, of ‘Pinot noir’ grapes, compared to berries from adjacent vines free of GLRaVs (same vineyards). Three different rootstock/scion combinations were included in this study. Clusters were collected for two growing seasons from commercial vineyards in the Willamette Valley of Oregon, and each vine sampled was tested for GLRaV-1, -2, -3 and Rupestris stem pitting-associated virus (RSPaV). All sampled vines were infected with RSPaV. Grapevine leafroll associated virus -infected vines tested positive for GLRaV-2 or GLRaV-3. Overall, fruit infected with GLRaV-2 and -3 had reduced percent soluble solids, decreased individual and total anthocyanins, and increased skin and pulp weight for all three ‘Pinot noir’ rootstock/scion combinations examined. Vitis riparia rootstock/‘Pinot noir’ clone 114 scion combination appeared to be the most sensitive to GLRaV-3 infection, having significant reduction of all five anthocyanins, total phenolics, and total tannins, with an increased cluster weight and 100-berry weight. No clear trends were observed in the polyphenolics analysed.

Nonhost Versus Host Resistance to the Grapevine Downy Mildew, Plasmopara viticola, Studied at the Tissue Level

Following inoculation of host and nonhost plants with Plasmopara viticola, the grapevine downy mildew, a histological survey was undertaken to identify the stage where its development is contained in nonhosts and in resistant host plants. Three herbaceous nonhost species, Beta vulgaris, Lactuca sativa, and Capsicum annuum, and three grapevine species displaying different level of resistance (Vitis vinifera [susceptible], Vitis riparia [partially resistant] and Muscadinia rotundifolia [totally resistant]) where inoculated by P. viticola using a controlled leaf disk inoculation bioassay. During the early steps of infection, defined as encystment of zoospores on stomata, penetration of the germ tube, and production of the vesicle with the primary hypha, there was no evidence of a clear-cut preference to grapevine tissues that could attest to host specificity. The main difference between host grapevine species and nonhosts was observed during the haustorium formation stage. In nonhost tissues, the infection was stopped by cell wall-associated defense responses before any mature haustorium could appear. Defense responses in resistant grapevines were triggered when haustoria were fully visible and corresponded to hypersensitive responses. These observations illustrate that, for P. viticola, haustorium formation is not only a key stage for the establishment of biotrophy but also for the host specificity and the recognition by grapevine resistance factors.

Identification, isolation and characterization of a CC-NBS-LRR candidate disease resistance gene family in grapevine

Plasmopara viticola causes downy mildew of grapevine, one of the most important diseases in viticulture. Resistance to this oomycete is present in American and Asian Vitis species, while traditional European Vitis vinifera cvs. for wine and table grape production are susceptible. Breeding aims to achieve resistance through introgression, but the molecular mechanisms are still unknown. Therefore, the differential display approach was used to detect grapevine genes involved in defense. P. viticola sporangia were applied to the lower leaf surface of in vitro plants of the resistant Vitis riparia selection ‘Gloire de Montpellier’ and susceptible cv. ‘Riesling’. Controls were treated with sterile water. Messenger RNAs extracted 12 h post infection were subjected to differential display. Seven transcripts appeared specifically induced during the incompatible interaction. Sequencing showed that they build three classes. One of them, named VRP1, represented by three transcripts of almost identical sequence but differing lengths, showed clear homology to resistance genes of the NBS-LRR type from other plants. Northern hybridizations confirmed its elevated expression in the resistant ‘Gloire de Montpellier’. Redundancy of VRP1 PCR products from V. riparia prevented PCR-walking, so the VRP1 genes were isolated from a BAC-library of the resistant cv. ‘Regent’. Three genes matching the original VRP1 sequences were found within a BAC clone carrying a 134,392 bp insertion. These were referred to as VRP1-1, 1-2 and 1-3. They encode proteins of 798, 811 resp. 813 amino acids and exhibit the structure of CC-NBS-LRR resistance genes. They were genetically mapped to linkage group 10.

Use of in vitro method to evaluate some grapevine varieties for tolerance and susceptibility to sodium bicarbonate-induced chlorosis

ABSTRACT Nodal shoot segments of four grapevine genotypes well known for their Fe-chlorosis characteristic [Vitis vinifera × Vitis berlandieri Fercal, resistant; V. berlandieri × Vitis rupestris 1103 P, mid-resistant; Solonis (Vitis riparia × V. rupestris × Vitis candicans) × Othello (Vitis labrusca × V. riparia × V. vinifera) 1613 C, susceptible; V. vinifera L. cv. Perlette, resistant] were cultured in vitro. The effects of three levels of iron sodium ethylene-diaminotetraacetate (FeNaEDTA; 9, 18, 36 mg l−1) and three mixtures of iron and 840 mg l−1 NaHCO3 (sodium bicarbonate) in the Murashige and Skoog (MS) medium supplemented 4.9 μM indole-3-butyric acid (IBA) were compared. We assayed the chlorosis rating of leaves, total chlorophyll of leaves, dry shoot weights of the plantlets, and active and total Fe content of the leaves. The most suitable concentration in determining the reaction of genotypes to iron chlorosis was 9 mg l−1 FeNaEDTA. Bicarbonate addition had negative effects on the iron intake and...

The Interaction between Soil Nutrients and Leaf Loss during Early Establishment in Plant Invasion

Abstract Nitrogen availability affects both plant growth and the preferences of herbivores. We hypothesized that an interaction between these two factors could affect the early establishment of native and exotic species differently, promoting invasion in natural systems. Taxonomically paired native and invasive species (Acer platanoides, Acer rubrum, Lonicera maackii, Diervilla lonicera, Celastrus orbiculata, Celastrus scandens, Elaeagnus umbellata, Ceanothus americanus, Ampelopsis brevipedunculata, and Vitis riparia) were grown in relatively high-resource (hardwood forests) and low-resource (pine barrens) communities on Long Island, New York, for a period of 3 months. Plants were grown in ambient and nitrogen-enhanced conditions in both communities. Nitrogen additions produced an average 12% initial increase in leaf number of all plants. By the end of the experiment, invasive species outperformed native species in nitrogen-enhanced plots in hardwood forests, where all plants experienced increased damage relative to control plots. Native species experienced higher overall amounts of damage in hardwood forests, losing, on average, 45% more leaves than exotic species, and only native species experienced a decline in growth rates (32% compared with controls). In contrast, in pine barrens, there were no differences in damage and no differences in performance between native and invasive plants. Our results suggest that unequal damage by natural enemies may play a role in determining community composition by shifting the competitive advantage to exotic species in nitrogen-enhanced environments.

Mercurial inhibition of root hydraulic conductance in Vitis spp. rootstocks under water stress

This work aims to quantify the contribution to whole-root water transport of the fraction controlled by cellular metabolism in grape rootstocks upon water stress. We used mercuric chloride as inhibitor of cell metabolism on genotypes obtained from hybridization of Vitis berlandieri with either Vitis rupestris or Vitis riparia, and we found that the fraction of root water transport under metabolic control is higher in former, which are known to be more resistant to water stress. In addition, as these rootstocks showed lower vessel embolization during water stress, we suggested a possible role of cellular metabolism on the control of root embolism.

Importance of internal hydraulic redistribution for prolonging the lifespan of roots in dry soil

Redistribution of water within plants could mitigate drought stress of roots in zones of low soil moisture. Plant internal redistribution of water from regions of high soil moisture to roots in dry soil occurs during periods of low evaporative demand. Using minirhizotrons, we observed similar lifespans of roots in wet and dry soil for the grapevine 'Merlot' (Vitis vinifera) on the rootstock 101-14 Millardet de Gramanet (Vitis riparia x Vitis rupestris) in a Napa County, California vineyard. We hypothesized that hydraulic redistribution would prevent an appreciable reduction in root water potential and would contribute to prolonged root survivorship in dry soil zones. In a greenhouse study that tested this hypothesis, grapevine root systems were divided using split pots and were grown for 6 months. With thermocouple psychrometers, we measured water potentials of roots of the same plant in both wet and dry soil under three treatments: control (C), 24 h light + supplemental water (LW) and 24 h light only (L). Similar to the field results, roots in the dry side of split pots had similar survivorship as roots in the wet side of the split pots (P = 0.136) in the C treatment. In contrast, reduced root survivorship was directly associated with plants in which hydraulic redistribution was experimentally reduced by 24 h light. Dry-side roots of plants in the LW treatment lived half as long as the roots in the wet soil despite being provided with supplemental water (P < 0.0004). Additionally, pre-dawn water potentials of roots in dry soil under 24 h of illumination (L and LW) exhibited values nearly twice as negative as those of C plants (P = 0.034). Estimates of root membrane integrity using electrolyte leakage were consistent with patterns of root survivorship. Plants in which nocturnal hydraulic redistribution was reduced exhibited more than twice the amount of electrolyte leakage in dry roots compared to those in wet soil of the same plant. Our study demonstrates that besides a number of ecological advantages to protecting tissues against desiccation, internal hydraulic redistribution of water is a mechanism consistent with extended root survivorship in dry soils.

In vitro multiplication and hardening of grapevine plants in aeriated media

In vitro cultures have widely been used in horticulture for rapid multiplication of new varieties and clones as well as to produce pathogen-free stock material. To improve efficient hardening and transfer in vitro grown grapevine plants were multiplied by cutting them into single-node internodes with the whole leaf. Microcuttings including the shoot tips were rooted in granulated perlite moisted with tapwater under sterile conditions. After 2-3 weeks the rooted microcuttings were supplied by nutrients and hardened by gradual opening and finally by complete removal of the lids of jars or plastic boxes used for growth. Using this method microcuttings of Vitis vinifera cvs. „Chardonnay", „Cabernet franc", „Riesling" and „Sauvignon blanc" and the rootstock varieties Vitis riparia x Vitis cinerea cv. „Barrier" and Vitis berlandieri x Vitis rupestris cv. „Richter 110" formed new roots and shoots and 100% of the tested plants survived the acclimatization procedure. Similar results were obtained when perlite was replaced with rockwool-, or pit-pot blocks. This method may highly increase the efficiency of producing pathogen-free propagating material and new transgenic lines.

CBF4 is a unique member of the CBF transcription factor family of Vitis vinifera and Vitis riparia

The CBF/DREB1 transcription factors control an important pathway for increased freezing and drought tolerance in plants. We report here the isolation of one CBF/DREB1-like gene, CBF4, from both freezing-tolerant wild grape (Vitis riparia) and freezing-sensitive cultivated grape (Vitis vinifera). The deduced protein in V. riparia is 99% identical to the corresponding protein in V. vinifera; 45-48% to three other Vitis CBF proteins reported earlier and 57% to AtCBF1, and contains CBF-specific amino acid motifs. Agroinfiltration experiments in tobacco leaves revealed that VrCBF4 activates expression from reporter genes driven by a CRT-containing promoter. Expression of the endogenous Vitis CBF4 genes was low at ambient temperature, but enhanced upon exposure to low temperature (4 degrees C). Uncommon for CBF genes, this expression was maintained for several days. No significant difference in expression pattern was observed between V. riparia and V. vinifera. Vitis CBF4 was expressed in both young and mature tissue, in contrast to the previously described Vitis CBF1, 2 and 3. Together, these results suggest that CBF4 represents a second type of CBF in grape that might be more important for the over-wintering of grape plants.

Morphological and anatomical development in the Vitaceae. X. Comparative ontogeny and phylogenetic implications ofCissus quadrangularisL.

Although the grape family, Vitaceae, is of great economic importance, there is little understanding of the relationships between and within the genera. Recent DNA-based phylogenies of the Vitaceae have disagreed with traditional classifications by grouping some Cissus species (e.g., Cissus antarctica Vent.) in a clade with Vitis species, while placing other Cissus species (e.g., Cissus quadrangularis L.) in a monophyletic clade. This study investigates Cissus phylogeny using epi-illumination light microscopy and histological ontogenetic comparisons of vegetative and reproductive characters of Vitis rotundifolia (Forssk.) Vahl, Vitis ‘Ventura’, Vitis riparia Michx., C. antarctica, and C. quadrangularis. Ten characters limited to the Cissus species (connection of stipules, precocious axillary bud initiation, prophyll growth not obviously unequal, timing of mature shoot architecture on axillary buds, lack of spiral inflorescence branching, absence of calyptra, petals closely enveloping anthers, large gynoecial disk, ovary buried in the disk, and production of copious amounts of nectar) support the traditional phylogenies: the monophyly of Vitis and the exclusion of Cissus species from Vitis. The comparison of C. antarctica (Vitis–Cissus clade) and C. quadrangularis (monophyletic Cissus clade) supports the division of Cissus into two clades based on supernumerary buds, stipule connectivity, uncommitted primordium shape, and inflorescence branching patterns.

Morphological and anatomical development in the Vitaceae. IX. Comparative ontogeny and phylogenetic implications ofVitis rotundifoliaMichx.

The grape family, Vitaceae, is an economically important fruit family. Although the Vitaceae have been used for conservation, viticulture, and horticulture, little is known about the phylogenetic relationships of this family. Recent phylogenetic reconstructions using DNA data have disagreed with traditional classifications and other DNA-based phylogenies by grouping Vitis and Cissus species within one clade and linking Vitis rotundifolia Michx. (muscadine grape) more closely with these Cissus species. This study uses comparative developmental morphology as an independent method to investigate whether V. rotundifolia is more similar to Cissus antarctica Vent. (kangaroo vine) than other Vitis species. Results obtained using epi-illumination light microscopy and histology were compared with those from similar studies of Vitis ‘Ventura’, Vitis riparia Michx., and C. antarctica. Twelve vegetative and floral characters including axillary bud dorsiventrality, uncommitted primordium shape, calyptra, and ring-shaped gynoecial disk placed V. rotundifolia in with other Vitis species. Only two characters, stipule timing and simple tendrils, were common to C. antarctica and V. rotundifolia, suggesting that V. rotundifolia is more similar to Vitis spp. than to C. antarctica, and supporting traditional classifications contrary to ITS1 and trnL DNA phylogenies.