1. Introduction
Grapevines belong to the genus
Vitis L., which comprises about seventy species of lianes in the family Vitaceae Juss. Grape (
Vitis vinifera L. ssp.
vinifera) is one of the most ancient fruit-bearing plants in the Old World, and together with the olive, fig, pomegranate and date palm, it comprises the oldest groups of domesticated fruit trees [
1,
2,
3]. Today, it is widely accepted that the modern cultivars of
V. vinifera are the result of the domestication of
V. vinifera ssp.
sylvestris (C.C.Gmel.) Hegi (wild grape) [
4,
5,
6]. The main feature used to distinguish between wild and cultivated
Vitis is the reproductive system: the wild grape is dioecious, whereas the cultivated grapevine has hermaphroditic flowers; however, a small percentage of hermaphroditism also exists in the wild grape populations [
7]. The domestication process, likely involving vegetative propagation of clones by growing cuttings, induced important changes in the reproductive biology of the grape cultivars, resulting in different properties; both physical (size, color), nutritional (sugar, carbohydrate and phenolic compounds), as well as in seed shape diversification. Ampelography, the botanical discipline dedicated to the identification of grapevines, has been traditionally focused on the shape of vine leaves and grape berries, and it has more recently been focused on DNA fingerprinting [
8]. The seeds of grape, often called pips, are highly polymorphic and have an important role for the taxonomic study within the genus
Vitis [
9,
10,
11,
12,
13,
14,
15]. The description of seed shape proposed in this work is complementary to morphometric analysis and can be useful in Ampelography.
In early morphological studies with grape seeds, two morphotypes were defined consisting of pear-shaped or oblong to ovoid seeds, that corresponded respectively to the taxonomic groups Euvitis and Muscadinia [
9,
10]. The seeds of cultivated species (
V. vinifera,
V. labrusca L.,
V. aestivalis Michx.) are larger and their dimensions are more variable than those of wild species [
9,
10,
11]. Stummer’s index was proposed to discriminate between cultivars and wild forms, defining the “wild syndrome” for those seeds having values of breadth/length above 0.8 and corresponding to wild extra-European
Vitis species [
9,
12]. The seeds of cultivated grapevines have, in general, elongated, pear-shaped seeds, with a long beak, while the wild forms have shorter seeds, although caution must be taken in the identification of the latter, because of the difficulty in differentiating wild forms and feral cultivars [
9].
Seed morphology departs from two points: first, it requires a concise description, based in qualitative aspects; and, second but not less important, metrology, i.e., the set of magnitudes that provide the dimensions of a seed. Metrology may also confirm, define and expand information about the adjectives used in the former, qualitative, description. Adjectives used to describe seed shape in
Vitis include some already mentioned, like oblong, ovoid or pear-shaped (pyriform) and others like cordate, globose, oval, rounded, squat or triangular [
10,
11,
13]. Magnitudes used to measure seed shape include seed length, seed breadth, chalazal length, chalazal position and others describing primary aspects of the morphology of pips, as indicated by Jaquat and Martinolli [
1] and Mangafa and Kotsakis [
13]. Modern methods of automated vision are based on the coordinates of the points in a bi-dimensional image and permit new approaches to seed morphology. Digital image analysis coupled with morphometric studies of grape seeds is a relatively new application in the plant research field. By using a set of high-resolution digital images, it is possible to obtain measurements of morphometric, colorimetric and textural features, describing the shape, size, color and texture of seeds, and this quantitative information can be correlated with various qualitative aspects of the seeds. The data obtained, corresponding to magnitudes such as area, perimeter, circularity and roundness [
13,
14,
15,
16,
17], can be combined and subjected to analysis based on diverse algorithms, for example, elliptic Fourier (EFD) and Aralik descriptors. Finally, in all cases, the data are collected in a matrix and can be submitted to Linear Discriminant Analysis (LDA), to classify the seeds in groups [
9,
14,
15,
18,
19,
20,
21].
There is an interest, both theoretical and applied, in joining both aspects of morphology: the descriptive adjectives and the measurements with the objective of testing if the groups defined by artificial vision and statistical analysis correspond to well defined geometric figures. This objective, that we may define as obtaining new measurable adjectives, can be reached by comparing the seed images with geometric models based on mathematical curves. This way, we obtain the J index that gives the percent of similarity between the two images (the bi-dimensional image of the seed and the geometric model). High values of J index were obtained when comparing images of seeds with the cardioid or modified cardioids in the model plant
Arabidopsis thaliana (L.) Heynh [
22], as well as in the model legumes,
Lotus japonicus L. and
Medicago truncatula Gaertn. [
23], also in
Capparis spinosa L. [
24], and in species of the Papaveraceae and Malvaceae [
25,
26].
Other geometric models include the ovoid and the ellipse, that give high values of J index with seeds of species in the Cucurbitaceae [
27] and in the Euphorbiaceae, such as
Ricinus L. and
Jatropha L. [
28,
29]. Seed shape in wheat kernels was described based on three geometric figures: (1) an ellipse of aspect ratio (AR) = 1.8 fitting the “round varieties” (
T. aestivum L. ssp.
aestivum cv. Zebra and Torka), (2) a lens of AR = 3.2 for the elongated kernels (
T. monococcum L.), and (3) an ellipse of AR = 2.4 that adjusted well the kernels of intermediate-shaped varieties such as
T. durum Desf. cv. Floradur [
30]. This latter work, demonstrating the adjustment of seed shape to geometrical forms, is useful to complement the results of morphometric analysis as those reported in
Vitis sp. [
9,
14,
15,
18,
19,
20,
21].
Our objective was to describe seed shape in species, subspecies and cultivars of
Vitis by a combined geometric and statistical approach. The images of grape pips belonging to different genotypes were compared with geometrical models derived from mathematical curves that represent diverse heart shapes [
31,
32] or the pyriform curve [
33]. This allows the description of seed shape with adjectives that may be measurable. For example, cordate means heart-like, thus defining a family of curves, responding to different equations that include different models. Among the cordate figures (heart-shaped), the cardioid is the curve defined by the trajectory of the point in a circle rolling around another circle of the same radius. This curve responds to an algebraic equation and is reproducible, thus, for any given two-dimensional figure well defined by a profile, we may calculate the percentage similarity to the cardioid. News models based on geometrical figures are defined for seed shape quantification in the species and cultivars of
Vitis. In addition, the overall morphology of
Cissus and
Parthenocissus, genus related to
Vitis in the Vitaceae [
34] is described, presenting an overview of seed shape in the Vitaceae.
4. Discussion
Seven geometric models have been described for the first time and applied to the morphological description of seeds in the Vitaceae. M1, a heart curve defined by Weinstein [
31], is useful for the quantification of seed shape in species of
Vitis and for the differentiation between the species
V. amurensis,
V. labrusca and
V. rupestris, in one side, and
V. vinifera, in the other. M1 adjusts better to seeds of
V. amurensis, V. labrusca and
V. rupestris than
V. vinifera. M6, the Fibonacci’s pear, adjusts well to seeds of
V. vinifera, in general, and better to
V. vinifera ssp.
vinifera than to
V. vinifera ssp.
sylvestris. This way,
V. vinifera ssp.
sylvestris can be differentiated from
V. vinifera ssp.
vinifera, because the values of J index obtained with M6 (the Fibonacci’s pear) are higher in the former. The water drop, a rounded heart curve (M3) corresponds better to the rounded seeds of species
V. amurensis and
V. rupestris than those of
V. labrusca, also giving high values of J index with the seeds of
Vitis vinifera ssp.
sylvestris. Thus, M3 permits to differentiate between
V. vinifera ssp.
vinifera and
V. vinifera ssp.
sylvestris, the former giving higher values of J index with this model. The combination of M3, M6 and M7 is useful for the differentiation of
V. vinifera ssp.
sylvestris from other species of
Vitis.
The models proposed are related to previous seed shape descriptions of the Euvitis and Muscadinia taxonomic groups. Models 1, 2 and 3 resemble the Euvitis type, while Models 4, 5, 6 and 7 fit better the Muscadinia shape [
9,
10]. The seeds of wild species are smaller, robust and with a rounded outline resulting in a heart-like shape, often termed as “cordate”, with short stalks and a flat ventral side with sharp angles and a strongly developed chalaza, while those of cultivars are large, elongated, oval or pyriform, with an elongated stalk [
9].
The identification of geometric curves resembling the outline of seed images complements the results of morphometric analysis and allows more exactitude in the description of species and cultivars and for their comparison. Among the two subspecies of grape, subspecies
sylvestris adjusts better to M3, while subspecies
vinifera adjusts better to M6, thus concerning seed shape,
V. vinifera ssp.
sylvestris resembles more other species of
Vitis than
V. vinifera ssp.
vinifera. This demonstrates a great capacity of variation in the shape of seeds of cultivars, where significant changes in seed shape are due to selection during cultivation [
28].
Concerning cultivars, Meserguera gave higher values with the elongated heart curve (M5), while Malvasia seeds adjusted better to M7. New models based on geometrical curves can be useful for the description of seed shape in other cultivars.
Seeds of Cissus verticillata adjust well to M2, while seeds of the species Parthenocissus tricuspidata adjust better to M4. The quantification of seed shape with M4 shows differences between seeds depending on the number of seeds per fruit. Fruits with one or two seeds have their seeds more rounded and with higher values of J index than the seeds from fruits with three seeds per fruit.
Both C. verticillata and P. tricuspidata seeds resemble more the seeds of V. amurensis, V. labrusca and V. rupestris than those of cultivars of V. vinifera, suggesting that the primitive seed type, associated with a reduced number of seed per fruit, resembles more Model 1.
The seeds in the Vitaceae present a very peculiar shape, for which there is no adaptation to a single geometric model, like the ellipse, the ovoid or the cardioid [
30,
32], but to more complex figures. This is in contrast with other families, where the ellipse, the ovoid or the cardioid are the models defining seed shape for most species and genera [
22,
23,
24,
25,
26,
27,
28,
29,
30,
32].
The magnitudes used in morphology have different information about the shape of a plane figure. Magnitudes such as the area, perimeter, length or width do not give much information on shape, and thus, are of little value on their own to discriminate between cultivars or species. Other measures, such as circularity or roundness are more informative when they are close to the unity, but their usefulness depends on the similarity of the figure with the circle. If the figure is different to a circle, low values are of no informative value. In contrast, J index compares the image of a plane figure with a similar geometric figure selected as a model. In the case of grape seeds, J index is a way to provide valuable information in a single measure and, in consequence, useful in ampelography studies.
The current edition of the OIV Descriptor List for Grape Varieties and
Vitis species [
39] concerning seeds includes only three variables: length, weight and the presence of transversal ridges on dorsal side of seeds. J index is a new way of describing seeds that allows one to relate the variety studied to its genetic origin. The longer seed varieties are of eastern origin and generally table grapes, as seen in the descriptor, and the varieties of western origin are shorter and rounded linked to the origin of the domestication of most wild vines, as, for example, varieties of French and Spanish origin [
40,
41]. The work described in this article allows an approximation to elucidate if the variety is of eastern or western origin. Intermediate varieties such as Pinot and Merlot are of mixed origin. Further study of Spanish varieties where there is an Arabic influence would allow us to test this methodology.
The description of seed images by comparison with geometric figures will contribute to the validation of results obtained by automated artificial vision methods, improving the description and identification of species and varieties. It is a new, low-cost and relatively simple technique that can help growers, producers or breeders in seed identification.