Ampelometric and Ampelographic Characterization of Leaves of Indigenous “Vitis vinifera ssp. Vinifera” in the North of Morocco ^†

Abstract

Morocco, with its Mediterranean climate and its various potentialities, conceals an important space for the extension of viticulture, in particular that of traditional grape varieties, which has undergone profound upheavals linked to economic, social and environmental constraints which have had a negative impact on genetic diversity. The present study aimed to evaluate the richness of the local phylogenetic heritage of the traditional vines in the North-West of Morocco. For this, we carried out a characterization of a collection of 1617 leaves, taken from 162 vine plants and belonging to 27 different traditional varieties. Thereafter, we used an ampelometric and ampelographic approach with the Super Ampelo software. The ANOVA test revealed the most discriminating parameters, which were the angles, the depth of the lateral sinuses in relation to the lengths of the ribs, and the relationships between all the parameters. In this sense, the qualitative parameters (OIV Codes) confirmed the presence of morphological diversity within the grape varieties studied; the study of general averages made it possible to specify the varieties with the large values of distance/angles and of ratio, and we have shown the presence of a great intra-varietal diversity in addition to that which is inter-varietal. The analysis in principal components allowed the grouping of the grape varieties into five groups according to their expressions by the quantitative parameters, and it confirmed the hypothesis of the influence of the external environment in addition to the gene pool on the grape varieties. This leads us to provide more efforts to maintain inter- and intra-varietal variability and to fight against genetic erosion and the threat of environmental changes.

1. Introduction

For centuries, man has not stopped domesticating animal or plant species. The vine is one of his great successes, a wild liana which over time has undergone very significant development and adaptation at the genetic, physiological and organoleptic levels, as well as varietal diversification [1]. Nowadays, most cultivated varieties are all of the Vitis vinifera species, representing more than 10,000 grape varieties [1,2]. Thus, the products of the vine would occupy the first economic place at the world level [3], explaining its place among the most cultivated fruit trees in the world with 7,528 million ha of area and a grape production of 691 million quintals [4].

Over time, the vine has shown a great capacity to develop on rich soils but also on steeply sloping, rocky and poor soils. The vine has always occupied an important place in the traditional Mediterranean landscape by its presence in these two wild and cultivated forms [5]. The culture of this species is well rooted in the traditions of the Maghreb peasant populations in general, and the Moroccan population in particular. However, the introduction of new wine and table grape varieties by globalization has generated a genetic erosion of the wine heritage [6,7]. This has led to a regression or even the disappearance of a large number of indigenous grape varieties that contain genes for adaptation and resistance to biotic and abiotic constraints.

Faced with the almost total ignorance of our wine heritage and the absence of a descriptive catalog of these grape varieties, it is imperative to recognize the local phylogenetic heritage, given the phytogenetic wealth that the country conceals in this area. In order to identify the diversity of traditional grape varieties in the traditional agroecosystems of the Rif Mountains and their possible conservation, we characterized the traditional grape varieties according to different approaches (ampelographic and ampelometric) recommended by the International Organization of the Vine and Wine (OIV).

2. Experiments

2.1. Sampling

The surveys took place in 13 traditional vineyards (Figure 1), located in the region of Tanger Tétouane Al-Hoceima. This is the northwest section of the Rif arc, which many authors refer to as “Jbala country”. Due to the energetic aspect of its relief, this ensemble is characterized as much by the originality of its environment (mountain, Mediterranean and Atlantic) as by the specificity of its role in both the ancient and contemporary history of the Kingdom. In this region, there are mainly four homogeneous zones: the Tangérois; the coast and the Mediterranean basins; the Bas Loukkos basin; and the Jbala, containing the sampling sites [8].

Sampling was carried out at the time of fruiting. In each prospected vineyard, with the help of the farmer, we collected samples of leaves for each named and recognized grape variety. Thus, for each plant, we collected 10 to 20 leaves located between the 7th and 12th node, counted from the base of the primary branch, according to the recommendations of the OIV [9].

2.2. In the Laboratory

After drying and organizing the leaves harvested in a herbarium, they were scanned and the images obtained were processed by the Super Ampelo measurement software, which is a program designed to help catalog the genetic material of the vine by ampelographic and ampelometric characters, according to the descriptors of the OIV [10].

3. Results

Although the area of distribution of the vine within the traditional agroecosystems of the Rif Mountains is very restrained, our study revealed a very important diversity made up of 27 traditional grape varieties: Ainab byed (AIB), Aineb Noir (AIN), Albaydi (ALB), Labyed (LBY), Alfasi (ALF), Alkouz (ALK), Babour Hmara (BABH), Boukhanzir (BKH), Boukwanes (BKW), Bezul laawda (BZA), Bezul laawda byed (BZAB), Bezul laawda hmer (BZAH), Chouikhi (CH), Douak (DAK), Eddibani (EDD), Feqqas (FKS), Hmar Bouaamar (HBA), Houmrani (HMR), Jenyani (JNY), Lechehab (LCH), Matecha (MTCH), Sbiyae Lebnat (SBLB), Taferyalt (TFR), Taferyalt byed (TFRB), Taferyalt Sfar (TFRJ), Taferyalt Lakhal (TFRN), Valenci (VAL).

3.1. Qualitative Characterization of the Adult Leaf

3.1.1. Shape of Blade

According to OIV 067, we distinguished between five groups of grape varieties (Figure 2); grape varieties with a pentagonal shape made up a proportion of 83%, while 8% had a wedge shape, 6% had a circular leaf, and only 1% had heart-shaped leaves.

3.1.2. Number of Lobes

According to OIV 068, the majority of grape varieties studied (96%) had leaves with five lobes, 2% with three lobes, 1% with seven lobes and 1% with one lobe (Figure 3).

3.1.3. Shape of Teeth

According to OIV 076, we noticed that the majority of grape varieties had rectilinear toothed leaves observed on both sides (99%). However, 1% had two-sided convex teeth (Figure 4).

3.1.4. Degree of Opening of Petiole Sinus

Concerning OIV 079, it was observed that on the sampled grape varieties, 57% presented leaves with a closed petiole sinus, while 31% had leaves with half overlapping sinuses and 12% had leaves with open petiolar sinuses (Figure 5).

3.1.5. Shape of Sinus

Taking into account the OIV 083 1, we noted the presence of two forms of sinus in the grape varieties studied, with 96% of individuals having U-shaped petiole sinuses, and 4% with V-shaped sinuses (Figure 6).

3.1.6. Teeth in the Petiole Sinus

Regarding OIV 081 1, it was noted that 94% of the grape varieties studied were characterized by the absence of teeth in the petiolar sinus, against 6% which had the particularity of the presence of teeth (Figure 7).

3.2. Quantitative Characterization of the Adult Leaf

The analysis of variance made it possible to specify the importance of the variability of the 86 ampelometric parameters of the leaves likely to differentiate the grape varieties. In this context, the analysis of variance for the 73 most discriminant parameters out of the 86 measured showed a highly significant difference between the different grape varieties (p <0.0001), which is the result of a significant morphological diversity within and between the different individuals that make up these grape varieties. This difference is expressed more with very high F for the characteristics related to the size and the base of the teeth (HD, BD), shape of the teeth (HBD1), petiole opening angle (PI), and surface (Area) (

Table 1. ANOVA analysis of the various ampelometric measured parameters.

Variable	Fisher’s F	Pr > F	Variable	Fisher’s F	Pr > F	Variable	Fisher’s F	Pr > F	Variable	Fisher’s F	Pr > F
HD1	185.648	<0.0001	ON41	6.595	<0.0001	ON1	5.042	<0.0001	ANGA1	3.319	<0.0001
BD1	44.433	<0.0001	LUPIC	6.206	<0.0001	OI	4.973	<0.0001	HN21	3.25	<0.0001
BD	34.627	<0.0001	O41N51	6.069	<0.0001	BN4	4.97	<0.0001	LULA	3.162	<0.0001
HBD1	16.126	<0.0001	O31N41	5.933	<0.0001	OS1	4.969	<0.0001	HD	3.032	<0.0001
RS1	10.574	<0.0001	FN2	5.746	<0.0001	OI1	4.895	<0.0001	AL1	2.995	<0.0001
RI	9.696	<0.0001	ON21	5.71	<0.0001	ON3	4.813	<0.0001	GA	2.981	<0.0001
RS	9.316	<0.0001	O4N5	5.684	<0.0001	HN2	4.64	<0.0001	ALBEOSOI1	2.935	<0.0001
PI	9.061	<0.0001	BN41	5.615	<0.0001	ALBEGA	4.232	<0.0001	GA1	2.909	<0.0001
BN2	8.962	<0.0001	BN21	5.574	<0.0001	ALBE	4.131	<0.0001	OO3	2.815	<0.0001
Area	8.043	<0.0001	ON2	5.463	<0.0001	TA	4.066	<0.0001	OO31	2.779	<0.0001
MU	7.622	<0.0001	O3N4	5.344	<0.0001	ALBEOSOI	4.041	<0.0001	ALBEGA1	2.707	<0.0001
RI1	7.612	<0.0001	HBD	5.262	<0.0001	SPSP1	3.942	<0.0001	OM	2.53	<0.0001
LAM	7.578	<0.0001	N4N41	5.242	<0.0001	ET	3.887	<0.0001	OM1	2.409	<0.0001
LUXLA	7.424	<0.0001	ON4	5.19	<0.0001	ANGA	3.878	<0.0001	BE	2.38	<0.0001
N2N21	7.393	<0.0001	ON31	5.171	<0.0001	TA1	3.826	<0.0001	ALBE1	2.121	<0.0001
LA	7.092	<0.0001	OS	5.124	<0.0001	AL	3.715	<0.0001	DE	2.059	<0.0001
LU	6.886	<0.0001	N3N31	5.067	<0.0001	HBN21	3.701	<0.0001	HBN4	1.659	0.001
OP	6.79	<0.0001	FN21	5.047	<0.0001	ET1	3.455	<0.0001	HBN2	1.503	0.009
BE1	1.336	0.047	R51	1.063	0.35	R5	0.965	0.553	R31	0.899	0.691
R21	1.087	0.305	R41	0.98	0.519	R2	0.909	0.671	RP	0.895	0.7
R4	0.895	0.699	R3	0.876	0.737	HN4	0.832	0.815	HN41	0.82	0.833
HBN41	0.65	0.982

).

4. Discussion

To get an idea of the structure of the variation in the various parameters measured, a principal component analysis was performed on a matrix encompassing the 27 traditional grape varieties, taking into account the 73 variables considered in the study (Figure 8).

Most of the variance expressed by axis F1 is explained by the correlation with the variables related to the following distances: leaf width (LA), leaf length (LU), length of veins and ribs (N1, N2, N3, N4 and N5), distance between vein tips (N2-N21, N3-N31 and N4-N41), leaf length including petiole (LUPIC), surface (Area), and petiole length (OP). As for axis F2, it is correlated with the variables related to the following angles: angle between I and I ‘(MU), angle between S and S1 (LAM), angle between N2 and N2’ (DE), angle between N1 and N2 (OM), angle between N2 and N3 (ET), angle between N3 and N4 (TA), the sum of the angles AL + BE (ALBE), and the sum of the angles AL1 and BE1 (ALBE1). This axis is also correlated with the ratio between the sum of the angles AL + BE and between the sum of the petiolar sinuses OS and OI (ALBEOSOI).

Considering the number of grape varieties studied, we carried out two projections; one made up solely of “Taferyalt”, a grape variety which is more representative of the sites; and a second projection, encompassing the rest of grape varieties without “Taferyalt”.

The PCA grouping together all varieties with the exception of “Taferyalt” (Figure 9) showed the existence of three categories depending on the variation of distances and angles. Category 1 constitutes the pole of the positive side of axis F1, which groups LBY (100%), VAL (100%), MATCH (100%), BZAB (100%), then CH (75%), FKS (60%), ALB (50%), and BZA (50%); this category is distinguished by large corner sheets. Category 2 constitutes the intermediate pole which includes HBA (100%), SBLB (100%), BKH (100%), ALF (100%), JNY (100%), DAW (100%), AIB (100%), LCH (100%), ALK (100%), FKS (40%), BZA (50%), ALB (50%), EDD (40%) and CH (25%); this category is distinguished by medium sized corner sheets. Category 3 constitutes the pole of the negative side of axis F1 which includes AIN (100%), BABH (100%), BZAH (100%), BKW (100%), HMR (100%), and EDD (60%); this category is distinguished by sheets of angles of small size.

However, the FKS, BZA, ALB and CH grape varieties are divided between categories 1 and 2, and the EDD grape variety between categories 2 and 3.

Regarding the second projection of the ACP with the “Taferyalet” grape varieties (Figure 10), we also made a distinction between three categories: Category 1 constitutes the pole of the positive side of axis F1, by grouping TFRN (100%) and TFRJ (100%); Category 2 constitutes the intermediate division regrouping TFR (100%); and Category 3 constitutes the pole of the negative side of axis F1, comprising TFRB (100%).

Although the study was based only on the morphological characters of the leaf, a significant intra- and inter-grape variability has been demonstrated. The interest and importance of this variability are particularly interesting insofar as they have been demonstrated over a small and relatively delimited territory.

5. Conclusions

This study constitutes a contribution to the characterization of the traditional grape varieties of the vines in the traditional agroecosystems of the Rif Mountains, using ampelographic and ampelometric parameters. These approaches are complementary and allow a better understanding of the diversity of local varieties of the vines. This study allowed us to collect preliminary data concerning the phenotypic characteristics of traditional grape varieties.

From the ampelographic approach, it emerges that the majority of the quantitative and qualitative parameters used demonstrate the existence of intra- and inter-grape variability within the individuals studied. As a result, it remains the main and essential tool for identifying grape varieties [11]. However, the morphological characters are influenced by environmental factors, such as soil properties [12], water availability and salinity [13], the nature of the rootstock [14], the level of nutrition [15], and the health of the plant [16], without forgetting the problem of synonymy and homonymy, especially in the case of vernacular names, which may differ for the same variety depending on the region of sampling. For this, it is of major importance to complete the present study with a molecular characterization.